
Class ft B^7 



Book - r J 



Copyright^ . 



COPYRIGHT DEPOSIT 




Author's Office Laboratory. 

For details and description see Appendix, page 274. 



ESSENTIALS 



OF 



LABORATORY DIAGNOSIS 



DESIGNED FOR 
STUDENTS AND PRACTITIONERS 



BY 

FRANCIS ASHLEY FAUGHT, M.D. 

DIRECTOR OF THE LABORATORY OF THE DEPARTMENT OF CLINICAL MEDICINE 

AND ASSISTANT TO THE PROFESSOR OF CLINICAL MEDICINE, 

MEDICO-CHIRURGICAL COLLEGE, ETC., ETC., 

PHILADELPHIA, PA. 



CONTAINING AN INDICAN SCALE IN COLORS, SIX FULL-PAGE 
PLATES AND NUMEROUS ENGRAVINGS IN THE TEXT 




PHILADELPHIA : 

F. A. DAVIS COMPANY, Publishers 

1909 



<G 









LIBRARY of CONGRESS 
Two Cooies Recerved 

FEb 11 1909 

Oopy»»K,nt fcntry 

SLAS3 A- XXO No, 

copy s. 



COPYRIGHT. 1909, 

BY 

F. A. DAVIS COMPANY. 



[Registered at Stationers' Hall, London, Eng.] 



Press of F. A. Davis Company, 

1914-16 Cherry Street. 

Philadelphia, Pa. 



PREFACE. 



A KNOWLEDGE of so many branches of medicine is required 
of the medical student of to-day, and as the time at his com- 
mand is comparatively limited, a manual embodying the essen- 
tials of the subject such as this aims to do cannot but prove 
useful. At the same time, it is believed to contain all infor- 
mation necessary to provide a working outline of clinical lab- 
oratory methods for the busy practitioner. 

The book is not intended to take the place of the many 
excellent and exhaustive text-books on Clinical Medicine, but 
rather to supplement them, by pointing out to the busy student 
and practitioner simple and reliable methods by which he may 
obtain the information desired, without unnecessary expendi- 
ture of valuable time upon difficult, tedious or untried methods. 

The author has endeavored in this work to present in as 
concise a manner as possible a selection of the analytical 
methods employed in the clinical laboratory, without burden- 
ing the reader with unnecessary detail, cumbersome methods, 
etc., many of which are extremely difficult, often requiring 
considerable knowledge of general chemistry and elaborate 
apparatus, which may place them beyond the reach of the 
practitioner. 

For these reasons it is hoped that this little work will 
prove equally valuable to the student and the practicing phy- 
sician. 

In preparing this work special effort has been made to 
bring the subjects treated up to date, by the introduction of 
such new methods as have proven reliable. Some of the 

(iii) 



i v PREFACE. 

material is entirely new, and many of the plates and cuts have 
been prepared from original drawings and photographs by the 
author. Special mention is to be made of the plate and draw- 
ings (Plate II, Figs. 17 and 18) of the normal relations of the 
stomach, which were prepared expressly for this work by Dr. 
George E. Pfahler. 

The appendix has been arranged to furnish a working basis 
for the equipment of a clinical laboratory, at the same time 
affording reference for the preparation of stains, reagents, etc., 
mentioned in the text. 

The leading authorities have been freely consulted, and 
much material has been obtained from such authorities as 
v. Jaksch, Sahli, Caille, Grawitz, Krehl, Max Braun, Tyson, 
Abbott, Purely, Eemsen, and Holland. 

The author takes this opportunity to express his appre- 
ciation of the many valuable suggestions received from Drs. 
Judson Daland and Wm. Egbert Eobertson, and Mr. Geo. B. 
Johnson, of the F. A. Davis Company; also, to his associate, 
Dr. Francis J. Dever, for invaluable aid in the preparation of 
the manuscript and in correcting the proof. 

F. A. F. 

5231 Baltimore Avenue, 
Philadelphia. 



INTRODUCTORY NOTE. 

By Judson Daland, M.D. 

Professor of Clinical Medicine, Medico-Chirurgical College. 
Philadelphia. 



This book should be in the possession of every medical 
student and most practicing physicians. It contains, as its 
title implies, the essentials, and only the essentials of those 
procedures necessary to clinical laboratory diagnosis. The 
peculiar value of this book resides in the concise and practical 
manner in which each subject is treated and each test 
described, and the entire absence of all superfluous data. 

This book is particularly well suited, not only to the med- 
ical student in the preparation of cases assigned to him for 
study, but also to practicing physicians, who have grasped the 
necessity of establishing a small laboratory in connection with 
every-day work. Those possessing such a laboratory will find 
it necessary to consult this book at short intervals. The more 
it is employed the more will its practical value be demonstrated. 
I unhesitatingly recommend this work to medical students, and 
trust that it will find a place in the laboratory of every prac- 
ticing physician. 



(v) 



LIST OF ILLUSTEATIONS. 



PAGE 

Author's Office Laboratory Frontispiece 

Plate I. Intestinal Parasites of Man. . 104 

Plate II. Skiagraph. Normal Stomach. Child, Age 4 Years 140 

Plate III. Indican Scale. ( Colored ) 180 

Plate IV. Calcium Oxalate Crystals and Phosphates 224 

Plate V. Uric Acid Crystals, Cholesterin, Cystin, Tyrosin and Leucin 224 
Plate VI. Uric Acid Crystals 224 

FIG. 

1 . American make Microscope, showing Triple Nosepiece, etc 2 

2. Mechanical Stage 8 

3. Maltwood Finder 10 

4. Bronchial Cast, from case of Fibrinous Bronchitis in service of 

Dr. Judson Daland (Original) 22 

5. Fleischl Hemoglobinometer 32 

6. Thoma-Zeiss Hemocytometer in case. (A. H. Thomas) 36 

7. Appearance of Field of Thoma-Zeiss Hemocytometer, when properly 

mounted for counting the red corpuscles 38 

8. Daland Hematokrit. (A. H. Thomas) 39 

9. Ruling of Chamber of Thoma-Zeiss Hemocytometer 42 

10. The Author's Sphygmomanometer. (Pilling & Co. ) 77 

11. The Author's Sphygmomanometer, showing method of packing' 

Arm-band and Bellows in case. (Pilling & Co. ) 78 

12. The Author's Sphygmomanometer, showing Cuff applied and 

ready for blood -pressure determination 80 

13. Wright's Coagulometer 91 

14. Bogg's Coagulometer, diagrammatic representation 93 

15. Outfit for Gastric Test-meal Removal, Lavage and Inflation 119 

16. Diagrammatic Representation of Arrangement of Bottle for Meas- 

uring Cubic Contents of Stomach 125 

17. Solid Line shows form and relation of typical normal Stomach, 

with subject in standing posture 139 

18. Dotted Line shows form and location of normal Stomach during 

filling. Shaded portion shows contraction and change in form 

of Stomach 140 

19. Strasburger Apparatus. (After Steele) 151 

20. Various Forms of Urinometers and Urinometer Cylinders 169 

21. Westphal Balance. (A. H. Thomas) 170 

22. Various Forms of Ureameters 187 

23. Urinary Sediment 190 

24. Esbach Albuminometer 199 

25. Einhorn Saccharimeter during Performance of Test 207 

26. Water-power Centrifuge 219 

27. Eleztric-power Centrifuge 219 

28. Hand-power Centrifuge 216 

29. Sedimentation Glasses 220 

30. Centrifuge Tubes 223 

31. Arnold Steam Sterilizer. (A. H. Thomas) 260 

32. Autoclave. (A. H. Thomas ) 261 

33. Thermostat or Incubator. ( A. H. Thomas ) 272 

34. Portable Urinalysis Set 275 

(vi) 



CONTENTS. 



PAGE 

Section I. The Microscope 1 

Selection of the Instrument, 1. Care of the Microscope, 1. The 
Parts of the Microscope, 2. The Oil-Immersion Objective, 4. 
Apochromatid Objectives, 5. Illumination, 6. Cleaning the Micro- 
scope, 7. The Mechanical Stage, 8. The Warm Stage, 9. The 
Maltwood Finder, 9. Examination of Urinary Sediments, 11. 
Examination of the Blood, 16. The Camera Lucida, 17. 

Section II. The Sputum 19 

General Considerations, 19. Gross Composition, 20. Color, 21. 
Reaction, 23. Air Content, 23. Odor, 24. Hemoptysis, 24. Macro- 
scopic Examination, 25. Microscopic Examination, 26. Prepara- 
tion of Stained Specimens, 29. 

Section III. The Blood 31 

Chemical Composition, 31. Methods of Obtaining Specimen, 31. 
Appearance of Fresh Blood, 32. Specific Gravity, 33. The 
Amount and Reaction, 34. Estimation of Per Cent, of Hemo- 
globin, 35. Estimation of Erythrocytes, 37. Hematokrit Deter- 
mination of Erythrocytes, 39. Percentage of Erythrocytes, 40. 
The Color-Index, 41. The Volume-Index, 41. Estimation of Leu- 
kocytes, 42. Examination of Fresh Drop, 43. Staining Methods, 
45. Hematologic Terms, 47. Varieties of Leukocytes, 48. Differ- 
ential Count, 49. Leukocytosis, 50. The Anemias, 52. Chlorosis, 
54. Leukemia, 54. Spectroscopic Examination, 57. Bacteriologic 
Examination, 58. Spirochete Pallida, 59. Agglutination Reactions 
— Widal Reaction, 61. 

Section IV . Opsonic Method 64 

The Theory, 64. Method of Determining Opsonin Index, 67. 
Standardization of Vaccines, 69. Therapeutic Application, 69. 
Diagnosis of Obscure Localized Infections, 71. Immunization in 
Mixed Infections, 72. The Nephelometer, 73. 

Section V. Blood-Pressure 76 

General Considerations, 76. Blood-Pressure and Heart-Power, 77. 
Arterial Blood-Pressure, 79. Sphygmomanometer, 79. Systolic 
Pressure, 80 and 82. Diastolic Pressure, 82. Pulse Pressure, 82. 
Variations and Blood-Pressure, 83. Clinical Value of Sphygmo- 
manometer, 85. Blood-Pressure in Disease, 86. 

Section VI. Coagulation 89 

General Considerations, 89. Wright's Method, 90. Method of 
Russell & Brodie, 93. Clinical Observations, 94. 

Section VII. Blood Parasites 96 

Plasmodium Malaria?, 96. Differential Table, 97. Detection of 
Organism, 97. Clinical Relations of Plasmodium, 98. Spirochete 
of Relapsing Fever, 99. Filaria Sanguinis Hominis, 99. 

Section VIII. Animal Parasites 101 

Definition, 101. Classification, 102. Protozoa, 103. Platyhel- 
minthes, 107. Nematodes, 113. 

Section IX. Determination of the Functions of the Stomach.. 118 

The Gastric Contents— Vomitus, 118. Methods of Obtaining Speci- 
men, 119. Preliminary Preparation of the Patient, 120. Modified 
Ewald Test-Breakfast, 121. Technic of Removal of Test-Meal. 
122. Inflation of the Stomach, 123. Determination of the Capacity 
of the Stomach, 125. Gastric Lavage, 126. The Acids of Diges- 
tion, 126. Estimation of Free Hvdrochloric Acid, 129. The Or- 
ganic Acids, 130. Test for Occult Bleeding, 131. Estimation of 
Peptic Activity, 132. Estimation of the Activity of Rennet, 136. 
Digestion of Starch and Sugar, 137. Test for Motor Functions, 
138. Rontgen Ray Examinations, 13S. 



(vii) 



yi ;i CONTENTS. 

PAGE 

Section X . The Feces 143 

Physical Characteristics, 143. Study of Intestinal Digestion, 144. 
Determination of Motor Functions of Gastro-Intestinal Tract, 146. 
Chemical Examination of Feces, 150. Occult Bleeding in Gastro- 
intestinal Tract, 153. Bacteria and Protozoa in Feces, 154. Clin- 
ical Significance of Examinations of Feces, 157. Foreign Bodies, 
Calculi and Concretions, 158. 

Section XI. The Urine 161 

General Considerations, 161. Description and Importance of the 
Urine, 163. Physical Characteristics of the Urine, 164. The 
Amount, 166. The Specific Gravity, 168. Estimating the Total 
Solids, 171. The Reaction, 172. Chemical Composition of the 
Urine, 174. Phosphates, 176. The Sulphates, 179. The Chlorides, 
180. The Organic Constituents of the Urine, 181. Urea, 182. Uric 
Acid, 187. The Urates, 189. Creatinin, 191. Oxalates, 192. Abnor- 
mal Constituents of the Urine, 194. Albumin, 194. Glucose, 201. 
Cammidge Reaction, 209. Acetone, 212. Bile Pigments, 215. 
Pyuria, 217. Epithelia in Urine, 218. Tube Casts, 218. Cylin- 
droids, 221. The Inorganic Sediment, 223. Urinary Concretions, 
225. The Diazo Reaction, 226. Drugs and Poisons in the Urine, 
227. 

Section XII. The Cerebro-Spinal Fluid 231 

The Presence of Cerebrospinal Fluid, 231. Determination of the 
Cell-Content, 232. Determination of the Proteid-Content, 234. In- 
terpretation of the Findings, 235. 

Section XIII. The Body Fluids 236 

Peritoneal Fluid, 236. Inflammatory Exudate, 236. Transudates, 
236. Pleural Fluid, 237. General Considerations, 237. Non-Inflam- 
matory Transudate, 237. Inflammatory Exudate, 237. Method of 
Making Permanent Stained Specimen, 237. Microscopic Appear- 
ance of Stained Specimens, 238. The Pericardial Fluid, 238. The 
Synovial Fluid, 238. Hydrocele Fluid, 239. 

Section XIV. Human Milk 240 

General Considerations, 240. Physical Characteristics and Com- 
position, 240. Examination of Milk, 241. Determination of the 
Fat, 241. Determination of the Sugar, 243. Estimation of the 
Proteids, 243. Chemical Composition, 243. 

Section XV . Bacteriologic Methods 244 

The Tubercle bacillus, 244. Differential Diagnosis, 246. Staining 
Methods, 246. Diplococcus Pneumonii. 248. Method of Staining 
Capsules, 249. Bacillus of Pfeiffer, 249. Bacillus Diphtheriae, 250. 
Gonococcus of Neisser, 231. Gram's Method of Staining, 252. Re- 
action of Bacteria to Gram's Method, 253. Loeffler's Method of 
Staining FJagella, 234. Sterilization, 255. Intermittent Steriliza- 
tion, 259. The Autoclave, 261. Chemical Sterilization and Disin- 
fection, 262. Preparation of Culture Media, 262. Nutrient Bouillon, 
263. Nutrient Gelatin, 265. Nutrient Agar-Agar, 266. Glycerin 
Agar, 266. Blood-Serum, 267. Loeffler's Blood-Serum, 269. Tech- 
nic for Plates and Petri Dishes, 270. The Incubator, 271. 

Section XVI. The Appendix 274 

Office Laboratory Cabinet, 274. Report Forms for Urinalysis, 
Sputum Examination, Blood Examination, Gastro-Analysis, Blood- 
Pressure and Feces Examination, 277-282. Table of Clinical Terms, 
283. Municipal Request Form for Widal Reaction, 284. Lists of 
Chemicals, Reagents and Apparatus, 285 to 299. Tables of Weights 
and Measures, 300-301. 



I. 

THE MICROSCOPE. 



SELECTION OF THE INSTRUMENT. 

In purchasing a microscope one should consider nothing 
but the best. This is true economy, for a cheap or inferior in- 
strument is always an unsatisfactory one which, when once 
bought, is hard to get rid of, and if later discarded for a better 
instrument, represents a total loss. The possession of a good 
microscope is an absolute essential. Its services are required in 
almost every investigation of modern clinical diagnosis, and 
there is hardly a chapter in this work which does not call for 
its use. While the finest and most expensive instruments are 
still of foreign make, there are, nevertheless, many of American 
make which compare favorably with the imported instruments. 
An example of a good American instrument is shown in Fig. 
1, and is one of a series manufactured by the Bausch and Lomb 
Optical Co. 

CARE OF THE MICROSCOPE. 

In handling any instrument it is necessary to see that it 
is always grasped by some one of its solid parts — the base or 
the standard. Some of the newer makes are provided with an 
aperture in the standard for this very purpose. 

If the instrument is to be in daily use it should be kept 
under a bell jar or in a specially prepared cabinet built above 
the work table. (See Frontispiece.) Eeference to Fig. 1 will 
illustrate the mechanism and the different parts with which one 
should become familiar before attempting to use the instrument. 
Efforts to use the microscope by any one unfamiliar with its 
different parts and adjustment should never be permitted, since 
careless handling may result in serious damage to the objective 
or other delicate parts. 

(i) 



THE MICROSCOPE. 



DESCRIPTION OF THE MICROSCOPE. 

Eeferring to Fig. 1, the various parts of the instrument 
about to be described may be located. 

The Ocular, or Eye-Piece, consists of one or more converg- 
ing lenses, the combined action of which is to magnify the 




Fig. 1.— American make Microscope showing Triple Nosepiece and 
Aperture in Standard to facilitate handling. 

image formed by the objective. This part is contained in a 
tube of its own which telescopes into the top of the barrel, and 
is the part which the eye approaches when viewing the object. 



DESCRIPTION OF THE MICROSCOPE. 

Various oculars are usually provided, and are numbered 
from 1 to 8. As the number advances the magnifying power 

of the ocular increases; at the same time the length of the 
tube decreases. It is advised when increased magnification is 
desired, to accomplish this by increasing the power of the ob- 
jective rather than by using stronger oculars. The clearness 
and definition of the picture will then be preserved, whereas 
if the power of the ocular is increased, it is usually at the 
expense of definition. 

It will be found of great convenience to cement into one 
of the oculars a fine hair or eye-lash. This should rest upon 
the upper surface of the lower lens of the ocular, and will always 
be in focus. This serves as a pointer, and can be used to single 
out from a blood or other specimen some special point of in- 
terest, so that it can be shown to another person. This is par- 
ticularly valuable in class demonstration. 

The Barrel, or Tube, is the large tube of the microscope 
which serves as a conductor of the rays as they pass from the 
objective to the ocular. 

The Objective consists of a system of converging lenses 
arranged at the lower end of the barrel, where it forms a mag- 
nified inverted image of the object. Upon this piece depends 
the magnifying power of the microscope. Most objectives are 
designated by numbers running from 1 to 15, these numbers 
representing the fractions of an inch at wmich the lens operates. 
Foreign makes are designated by letters which are not in any 
way directly comparable to the numbers of the American objec- 
tives. For one desiring three objectives, the most useful num- 
bers will be found to be the 3, the 7 or 6, and the 12-oil-im- 
mersion, the 3 and 6 or 7 being chiefly employed in blood-count- 
ing, while the 1.2th is necessary for blood-examination and bac- 
teriologic work. 

The Stage is the table fastened below T the barrel and in a 
right-angle plane to it. This serves to retain the object in a 
vertical plane to the optical axis of the instrument. The table 
is provided with spring clips to better hold the slide in position 
during examination. The newest Leitz scope is furnished with 
a movable (not mechanical) stage which, by the operation of 
two milled screws, is capable of slight movement in all direc- 



4 THE MICROSCOPE. 

tions. This, when a mechanical stage is not at hand, is of great 
service in searching blood and bacteriologic slides. 

The Reflector is a small mirror situated below the stage 
which serves to direct the rays of light upward through the 
object in the optical axis of the microscope. The reflector has 
two sides, one carrying a concave, and the other a plane, mirror. 

The Sub-Stage Condenser (Abbe's) consists of a system of 
lenses arranged between the stage and the reflector. The object 
of this part of the instrument is to collect and condense the 
rays as they come from the reflector, so that they are focused 
upon the object, thus furnishing brilliant illumination. 

The Iris Diaphragm is now universally employed for con- 
trolling the intensity of the illumination. This is held in the 
same carrier as the condenser, and is just below it. By means 
of a lever every gradation of light, from the most intense to 
absolute darkness, is readily obtained. The proper manipula- 
tion of this diaphragm constitutes a very important part of the 
practical knowledge gained from the use of the microscope, and 
has much to do with the success of many investigations. 

The Adjustments. — The coarse adjustment is the rack-and- 
pinion mechanism projecting from the upper part of the stand- 
ard, and is employed to rapidly raise and lower the barrel and 
its attachments. The fine adjustment is a micrometer screw 
situated usually below the rack and pinion. This serves the 
purpose of very gradually raising or lowering the barrel in order 
to obtain exact focus. 

The Draw-Tube is a very important adjunct of high-grade 
instruments, because by its skilful manipulation slight errors 
in refraction, due to inequalities in slide or cover-glass, may be 
corrected. 

The Nose-Piece, or collar, is fastened to the lower end of 
the barrel, and permits the attachment of two or three objectives 
at one time in such a position that, by rotation of the collar, 
any one of them may be immediately brought into the axis of 
the instrument. 

The most important accessories of the microscope are the 
objectives, and the quality of all microscope work largely de- 
pends upon their perfection. While the quality of objectives 
vary much, one cannot go far astray if they are obtained through 



APOCHROMATIC OBJECTIVES. 5 

a reputable supply house or from a well-known manufacturer, 
such as the Zeiss or Leitz abroad, and the Bausch and Lomb 
Optieal Co., V . S. A. 

THE OIL-IMMERSION OBJECTIVE. 

The oil-immersion objective, or homogeneous system, is so 
constructed that when in use the pencil of light passing through 
the object to the' objective traverses only media of the same 
refractive index. This is accomplished by placing between the 
cover-glass and the end of the objective a medium having the 
same refractive index as glass. To accomplish this a drop of 
cedar oil is placed upon the cover-glass, and the objective brought 
into contact with this, and the observation made through the oil. 
This class of lens is intended to work only with the oil, and is 
unsatisfactor} r when used dry. Frequently it is not convenient 
to use cedar oil for the preliminary examination often employed 
to determine the progress of staining, etc. ; here the staining 
fluid may be washed off and a drop of water used in place of 
the oil; this will give a sufficiently good picture for the pur- 
pose, and has the advantage of not interfering with the addition 
of further stain when desired. 

The tube or barrel of the microscope is made in two forms, 
the long or English type being from 8 to 10 inches, and the 
short or Continental type having a length of 6% inches. The 
latter is the more desirable form, since the majority of manu- 
facturers of objectives adjust them for the short tube. 

APOCHROMATIC OBJECTIVES. 

This term is applied to a particular variety of lens contain- 
ing a special kind of glass (containing calcium fluoride), be- 
sides the usual crown and flint glass, the object of this being 
to produce a greater degree of achromatism, thus reducing 
chromatic aberration. The special value of these objectives 
when used with a compensating ocular (compensating eye-pieces 
are specially constructed for use with apochromatic objectives), 
is as follows : Three rays of the colored spectrum, instead of 
two, as in case of the achromatic glasses, are focused in the 
same plane, leaving only a minute tertiary spectrum: also with 
these objectives the spherical aberration is corrected for two 



g THE MICROSCOPE. 

colors in the brightest part of the spectrum, and the objective 
shows the same correction for marginal rays as for the central 
part of the aperture. Finally, apochromatic objectives admit 
the use of highly magnifying oculars. This form of objective is 
particularly useful in photomicrographic work, where it is very 
important to abolish chromatic aberration. 

ILLUMINATION. 

Illumination constitutes a most important factor in the 
practical use of the microscope, and upon its proper manage- 
ment depends very much of the efficiency of the work. Direct 
unmodified sunlight is unsuited, and should not be employed 
for general work. North light is the most uniform and steady, 
and is to be preferred. The use of artificial light should, as far 
as possible, be avoided, and when it must be used its character 
and color may be greatly improved by inserting a piece of blue 
glass between the reflector and the object. It is always neces- 
sary to keep the eye close to the ocular in an endeavor to keep 
the unemployed eye open. Too much light is always to be 
avoided, as it is only an added strain, accomplishes nothing, and 
may materially interfere with the efficiency of the work. 

Oblique Light. — For urinary work the oblique light is best. 
By this term is meant light in which the parallel rays from the 
plane mirror meet the optical axis of the microscope at an angle. 
This form of light may be obtained in the following ways : (a) 
By placing the reflector to one side of the stage, this results 
in an oblique ray without materially lessening the strength of 
the light; the condenser must, of course, be swung out of the 
way. In microscopes which have a fixed mirror, i.e., only mov- 
able perpendicularly at right angles to the axis, oblique light 
may be obtained through the condenser, as follows : (b) First, 
focus the light upon the object through the condenser; then, 
lower the condenser until its focus is considerably below the 
plane of the object, thus the rays emerging from the condenser 
will decussate and then diverge, so that all but axial rays will 
fall upon the object in an oblique direction. 



TO CLEAN THE MICROSCOPE. 



TO CLEAN THE MICROSCOPE. 



Should the lenses or oculars become blurred or spotted from 
dust or dirt, proceed as follows: If the dirt is upon the ocular 
it will be discovered by rotating the ocular within the barrel 
while observing the illuminated field of the microscope. If the 
obstruction is upon the ocular it will be seen to move; if upon 
the objective it will remain stationary during this manipula- 
tion. To find an obstruction upon the objective rotate it, when 
the spot or mark will move with it. Finally, if after testing 
both the objective and ocular in the manner described above the 
location of the dirt or dust is not demonstrated, the trouble will 
then be found upon the glasses or spectacles if the observer 
happens to wear them. Having located the trouble, remove the 
affected part and cleanse as follows : If careful polishing with 
bibulous paper or fat-free silk fails to accomplish the desired 
result, then moisten the silk or paper with a trace of distilled 
water, using this to aid in the removal, finally polishing dry. 
For oily or resinous smears a small amount of alcohol may be 
used. Special care should then be exercised to avoid any excess 
of the solvent which might enter between the individual lenses 
and, dissolving the cement, ruin the part. After using the oil- 
immersion it should always be cleaned before leaving the instru- 
ment. After removing the excess of oil with the silk, wipe off 
the remainder with a trace of xylol or benzine, immediately 
wiping dry. The cover-glass preparation may be treated in a 
similar way. 

Glass surfaces should never be touched with the fingers, 
and great care should be exercised to avoid dropping and possi- 
ble fracture of the oculars and objective. All metal parts of 
the instrument should be kept free from liquids, particularly 
acids and alkalies, benzine, xylol, alcohol, turpentine, and 
chloroform. 

To clean the mechanical parts, none but the best machine 
oil should be used, and this only in small amounts and at long 
intervals. The polished brass requires nothing but occasion- 
ally polishing with clean chamois. 

The two keynotes to successful use and preservation of the 
microscope are : handle with care and keep scrupulously clean. 



g THE MICROSCOPE. 

THE MECHANICAL STAGE. 

Of the many aids to exact work in the realm of clinical 
medicine the mechanical stage is of great practicability and 
wide application. Every possessor of a microscope should aspire 
to the possession of a mechanical stage, for this mechanism is 
not only a great time saver, but will materially aid in the search 
for bacteria and pathologic cells in the blood, and is practically 
a necessity in making a differential count. 

Description. — Eeferring to Fig. 2, the general appearance 
of this instrument will be seen. It is designed for application 
to the stage of the microscope, upon which it is rigidly fastened 




Fig. 2.— Mechanical Stage. 

by a collar and set screw attached to the stand. It is important 
in this connection to note that most manufacturers of micro- 
scopes make a mechanical stage for their particular instrument, 
and which frequently is not interchangeable with microscopes 
of other manufacture, so that, before purchasing, one should be 
certain that the stage will fit the instrument for which it is 
intended. When attached to the stand the slide carrier should 
move in both directions without any undue force and without 
any irregularity or jarring of the mechanism, and to the limits 
of its movement should remain in close and even contact with 
the microscope stage. 

The mechanical stage is fitted with two milled screws 
which are movable in either direction, and which convey gradual 



THE MALTWOOl) FINDER. 9 

motion to the slide hold in the jaws of the instrument. One 
of these screws controls the vertical, and the other the hori- 
zontal, motion, so that by proper manipulation it is possible 
to rapidly and accurately go over the whole of a specimen. (For 
more detailed description of the use of the mechanical stage in 
the differential count, see chapter on Blood.) 

Most mechanical stages are provided with a millimeter 
scale and vernier reading to 0.1 of a millimeter, which serves 
not only to measure the size of small objects upon the stage, 
but is also very useful for locating objects of special interest 
upon any slide, so that they may again be found without dif- 
ficulty. (See below the Maltwood finder and Pepper's applica- 
tion without the mechanical stage.) 

THE WARM STAGE. 

A further differentiation and improvement upon the me- 
chanical stage is the stage prepared for preserving specimens at 
body-temperature during examination. This is provided with a 
thermometer which indicates the temperature of the object 
during examination. An extemporaneous warm stage has been 
described in another section (Blood Parasites) to wdiich the 
reader is referred. 

THE MALTWOOD FINDER. 

In microscopic work, especially in studying blood or bac- 
teriologic slides, some sort of "finder" is an essential part of the 
equipment. While the vernier scale, which is attached to most 
microscopes is fairly satisfactory for individual work, it does not 
fulfill all conditions demanded of it, since the same stage and 
microscope must always be used, and if by accident the relation 
between microscope and stage is altered ever so little, then all 
previous figures indicating location are rendered valueless. 

The Maltwood finder (Fig. 3) does not possess the above 
disadvantages, and can be used universally with uniform results. 

The Maltwood finder 1 consists of a heavy glass slide com- 
paring exactly in size with the ordinary microscope slide. The 
central third of this slide is covered with a close network of 



1 Wm. Pepper : Jour. Amer. Med. Assoc., July 20, 1908. 



10 



THE MICROSCOPE. 



intersecting rectilinear lines which form a large number of 
uniform squares. Each square contains two figures arranged 
one above the other, so that no two squares represent the same 
combination (see below). This marking has been placed upon 
the slide by a photographic process. 



1 

1 

2 

1 

3 

1 


1 
2 

2 
2 

3 

2 


1 
3 

2 
3 

3 
3 


1 
4 

2 
4 

3 

4 


1 

5 

2 

5 

3 

5 



All Maltwood finders are made interchangeable, the squares 
coinciding exactl)* in all slides. 

Method of Using the Finder. — If on looking over the slide 
with the mechanical stage a part of the field is discovered which 




^ \ v V V 

v\W\v 



\\\ \\ \\\\\Y V • V vav//^ -\V N A\ *\\v N\- 

v\\\\\ \ ^ N \V\\ ■ \^\\\v \A\\\\ ■•* >*\\Vn 



Fig. 3.— Maltwood Finder, showing Dr. Pepper's Brass Angle 
Substitute for Mechanical Stage. 

one desires to examine at a later time, the slide is carefully re- 
proved without altering the location of the mechanical stage 
and the Maltwood finder substituted; the square is then ob- 
served which corresponds with the center of the slide. This 
combination is to be jotted down, and then in order to make 
sure that the proper reading has been made, the finder and slide 
should be substituted for each other a number of times. Sup- 



EXAMINATION OF URINARY SEDIMENTS. H 



pose this reading was made 1 1? L then, when later this par- 
ticular field is sought the above process is reversed, and when 
the recorded square is in the center of the field, the slide is care- 
fully substituted, when the object sought will appear in the field. 

One of the chief advantages of the finder is that the slide 
may be sent to any one having a finder, with a note to look for 
such or such a square. For those not possessing a mechanical 
stage, Dr. W. Pepper has devised a small right angle of brass 
which ma}^ be easily carried in the pocket, and which may at 
any time be substituted for the mechanical stage, as the only 
purpose of the mechanical stage is to afford a fixed angle into 
which the slide and finder may be fitted. 

The illustration shows the arrangement of slide and brass 
angle, and illustrates the size of the right angle compared to 
the slide. The strip of brass from which the form is cut should 
be about one-sixteenth inch thick. 

EXAMINATION OF URINARY SEDIMENTS. 

Preparation of the Slide. — After concentration of the sedi- 
ment by centrifugation or sedimentation, the next point is to 
arrange this collection upon the slide for examination under 
the microscope. For this examination a few drops may be taken 
up in a pipette and allowed to fall upon the center of a clean 
slide, upon which a clean cover-glass is immediately placed. 
This will exclude dust, prevent rapid drying during examination, 
and will so flatten the field that frequent change in focus will 
not be necessary, while the different elements are more easily 
viewed and differentiated owing to their uniform separation and 
lack of overlapping. 

The proper removal of the sediment from the bottom of the 
container is an all-important part of this process, and upon 
this the successful finding of casts will frequently depend. This 
is best accomplished with a small nipple-pipette or section of 
narrow glass-tubing, one end of which is drawn out into a coarse 
capillary point. Having cleansed the slide and cover slip, ar- 
range them in a convenient place and then proceed to remove 
a few drops of sediment from the bottom of the tube. It is 
absolutely necessary, during this procedure, to prevent the en- 
trance of an excess of fluid into the pipette. This end is accom- 



12 THE MICROSCOPE. 

plished in the case of the pipette by carrying it clown into the 
fluid nearly to the sediment, then expressing a few bubbles of 
air before passing the tip beneath the sediment. With the 
straight tube the moist thumb or finger is held firmly upon the 
upper opening of the tube while its tip is carried to the bottom 
of the vessel, then by means of a slight rotation of the tube 
while relaxing the pressure above, a few drops of sediment are 
allowed to enter the tube. Firm closure of the upper end of 
the pipette is maintained while the tube is withdrawn and its 
contents transferred to the slide. To further guard against 
diluting the sediment, the outside of the tube should be wiped 
dry after it has been removed from the urine. 

Special Technic for Excessive Sediments of Phosphates, 
Pus, etc. — It is not uncommon to meet with urinary sediments 
of such volume and density that the grosser condition may com- 
pletely obscure the more important but less numerous elements, 
such as casts, pus, blood-cells, etc. In such cases the following 
technic is advised: After taking up the sediment deposit but 
one drop upon the slide; then from a clean pipette add a few 
drops of distilled water or, what is better, the clear urine from 
above the sediment. Agitate this till evenly mixed, and then 
apply the cover-glass and remove excess of fluid with strips of 
filter- or blotting-paper. Thus diluted not only is there better 
opportunity of finding casts, but the internal structure of the 
different elements will be much clearer. 

Microscopic Search. — After preparation of the specimen as 
above outlined, the slide is transferred to the horizontal stage 
and first viewed by low power and a rather subdued light. In 
this examination it is safer to begin with the objective below 
the focal point, then with the eye in position gradually rack 
upward, at the same time keeping the slide moving with the 
other hand. If this is continued carefully there will be little 
difficulty in discovering the objects in the sediment, even if few 
and small. Another advantage of this plan is that all danger 
of crushing the slide or fracturing the objective by forcibly 
racking down is eliminated, since the objective is never carried 
downward except when the relation of objective to slide is plainly 
seen. 

Having focused the sediment and regulated the light, the 



EXAMINATION OF URINARY SEDIMENTS. 13 

slide should be searched as follows: Slowly move the slide for- 
ward until one corner of the cover-glass appears in the field, 
then locate the lower right-hand corner and gradually move the 
slide upward in as straight a line as possible, until the opposite 
border is reached. Then move slightly to the left and proceed 
in a straight line downward to the lower margin again. This 
procedure is repeated until the whole area of the cover-glass 
has been searched. For those having a mechanical stage the 
slide may be placed in this, and the same movements carried 
out with its aid. In either case it is well to keep one hand upon 
the fine adjustment to sharpen up any obscure objects which 
may be encountered. 

Casts. — In searching for casts note any elongated struc- 
tures which may appear in the field, and study carefully their 
outline under varying conditions of light and focus. Note care- 
fully the fundamental characters of the body, whether granular 
or clear; if cellular elements are attached, such as epithelia, 
blood or pus, and if any of these are found whether they are 
well preserved as indicated by a sharp outline, or whether irregu- 
lar and ragged from disintegration and age. Note also if there 
are any small, round, highly refractive granules (fat globules). 
Observe the relative size of trie casts, as the predominance of 
certain sizes may indicate the location and character of the 
change in the kidneys. 

There will be little difficulty in finding granular or cellular 
casts, as they are highly refractive and stand out plainly. Not 
so with almost invisible hyaline casts, which are very difficult 
to find on account of their feeble refractive powers. As aids 
to the detection of hyaline casts, it is important to remember 
that their outline is often accentuated by oblique light, and that 
too bright a light may absolutely obscure them. 

Diagnosis of Casts. — The distinguishing features of casts 
under the microscope are: (a) Uniformity of marginal outline 
throughout the whole or greater part of their length, the edges 
stand out at all times as well defined borders which do not look 
accidental, but show the moulded effect of the kidney tubules. 
(b) For the most part casts are uniform in their individual 
diameters, and do not appear to suddenly bulge and then become 
as suddenly constricted. The body of the cast never appears 



14 



THE MICROSCOPE. 



split, though it may occasionally be found curved and rarely 
twisted, (c) Casts are rarely exceedingly long, being usually 
between three and eight times their diameter or width, (d) 
The ends of casts are either rounded like the finger-ends or they 
are abrupt and ragged, plainly giving evidence of fracture ; they 
never gradually taper off and disappear. 

Diagnosis of Cylixdroids. — These structures are very 
commonly met in the urine, and may become a source of more 
or less confusion and doubt. They are long, slender threads 
that have distinctive features which serve to distinguish them 
from casts. These distinguishing features are: (a) Their out- 
lines are always more or less irregular, indistinct, and non- 
linear, (b) Their diameters are consequently very variable. 
(c) Their ends often taper into long, slender points which may 
be split, bifurcated or branched, but they never possess the 
smooth, rounded or abruptly fractured terminations of casts. 
They are rarely ever cellular or granular, but under high power 
frequently present longitudinal striations extending through the 
whole or a part of their length, (d) They are usually very long 
and slender, their total length frequently exceeding the diameter 
of the field when viewed through the medium power, (e) They 
are usually bent and often twisted into grotesque forms. 

Epithelia. — Various forms of epithelia are found in 
nearly all specimens of urine, and careful note should be made 
of their form and size, as well as of the condition of their pro- 
toplasm and nuclei. They are highly refractive, and are there- 
fore plainly visible under the microscope, their outlines stand- 
ing out clear and distinct. The general character of the epi- 
thelia found corresponds to the usual three varieties found 
throughout the body, viz. : squamous or flat, cuboidal, and 
columnar or cylindrical. All epithelia. are more or less granular 
and possess one or more nuclei, though the latter are not always 
visible, having been disintegrated or become obscured by granu- 
lar degeneration of the protoplasm. Epithelia are subject to 
certain physical alterations when in the urine. By absorption 
of water they swell up and become more regular in outline, the 
small forms thus becoming spherical. The small cuboidal or 
columnar epithelia are most important, since they probably 
come from the tubules of the kidney. The larger varieties of 



EXAMINATION OF URINARY SEDIMENTS'. 15 

columnar and squamous cells come from the lower portion of 

the genito-urinary tract, the largest cells of all being the 
squamous vaginal epithelium. 

Crystals. — Little difficulty should be experienced in rec- 
ognizing and classifying the crystals found in the urine. For 
the most part they are comparatively large and highly refract- 
ing, and are therefore best viewed by moderate power and good 
illumination. One exception to this general rule occurs in the 
case of calcium oxalate, which may be very minute and exceed- 
ingly numerous. x\s a general rule, all crystals that show yellow 
or brown pigment are uric acid or urates, this characteristic 
serving readily to differentiate the group. The clear, large 
prisms -(coffin-lid) are triple phosphates, while the smaller forms 
of highly refractive stars, crosses and dumb-bells are calcium 
oxalate. Beference to the plates in the section on urinalysis 
will show the character of the common, as well as of the rare, 
forms of crystals occurring in the urine. 

Pus axd Blood-Cells. — For the satisfactory study of these 
small elements, a higher power objective than that used for the 
study of crystals and epithelia is required. Oblique light, with 
moderate illumination, will be found best. Occasionally great 
difficulty is experienced in differentiating blood, pus, and small, 
round epithelial cells. This confusion is not so likely to occur 
when all varieties are present in sufficient quantity in one speci- 
men to admit of careful comparison; but when only a few 
scattered cells of one variety are found, mistaken identity is 
quite possible. It is well, as a general guide, to bear in mind 
the relative size of these different elements. Pus corpuscles are 
the most common, and these should be thought of first. They 
are identified as small granular discs, having multi-nuclei. 
Somewhat smaller in size (about one-third) will be noted the 
pale, non-granular, non-nucleated discs which are the red blood- 
cells. While at least one-third larger than the pus cells, the so- 
called renal epithelia present a granular protoplasm with large 
nuclei. 

Finally, after determining the individual elements in a 
given specimen, it is important to note the degree of degenera- 
tion and the presence of fatty change, particularly, occurring in 
the casts; and, lastly, the number of each formed element per 



lg THE MICROSCOPE. 

field, or per drop, should be determined. This observation 
being important in following the course of any pathologic con- 
dition, as it is similar to the repeated quantitative determina- 
tion of albumin or sugar. 

Micro-organisms. — The presence of excessive numbers of 
micro-organisms, their motility, and even some of their mor- 
phologic characteristics may be determined during the micro- 
scopic examination of the sediment ; a complete and satisfactory 
examination of their detail and identity cannot, however, be 
determined except by special bacteriologic technic, for which the 
reader is referred to larger works on bacteriology or urinary 
diagnosis. 

EXAMINATION OF THE BLOOD. 

For an examination of fresh blood the 6 objective is most 
suited, since nothing but the cell outlines can be seen with the 3. 
A good light, with the condenser in close apposition with the 
under surface of the slide, will give the best results. To differ- 
entiate the varieties of white cells by their nuclei and granula- 
tion requires very careful regulation of the light, since the very 
slight difference in refraction of nuclei and protoplasm makes 
the differentiation extremely difficult at best. 

Counting the Corpuscles: Red Cells. — Having made the 
proper dilution and mounted the specimen as outlined in the 
section on blood, the slide is placed upon the stage and a few 
minutes allowed to pass while the corpuscles settle to the bot- 
tom of the chamber. The medium power, 6, is brought into 
close apposition with the cover-glass, and then with the eye to 
the ocular the fine adjustment will bring the individual cells 
into view. Considerable difficulty is at times experienced in 
locating the part of the counting chamber containing the ruled 
lines. This may be overcome by centering the inner circle of 
the chamber by means of the outside of the objective. If this 
is carefully clone the first attempt at focusing will usually bring 
the squares into view. Some hemocytometer slides are very 
faintly ruled, rendering the operation of counting very difficult. 
This may, in a measure, be overcome by reducing the light, at 
the same time taking advantage of the slight shadow caused by 
oblique rays coming from the lowered condenser. If this is done 



THE CAMERA LUCIDA. 17 

great care should be observed in determining border-line cells on 
account of the uncertain shadow cast by all colls with this 
illumination. 

The White Cells. — Either the 3 or the objective may he 

used in this connection, and of these the 3 is generally pre- 
ferred. It does, however, require a little more care and experi- 
ence to make an accurate count, hecause of the minute appear- 
ance of the individual cells. Its chief advantage is that it is 
not necessary to move the slide while counting the entire field, 
after it has once been centered, thus doing away with the error 
of missed squares which occasionally occurs in moving the 
slide when using the 6 objective. Here, again, low r illumination 
and oblique rays may advantageously be employed. 

Examination of the Stained Specimen. — Since the usual 
reason for staining and examining the dried specimen is to 
study minute structural characteristics, to make a differential 
count or to discover the presence of bacteria or parasites, it is 
necessary to employ the oil-immersion lens and to occasionally 
supplement this with a highly magnifying ocular. In this work 
good illumination and sharp definition (condenser close to 
slide) are essential. 

THE CAMERA LUCIDA. 

The camera lucida is a device by the aid of wdiich it is 
possible to quickly and accurately trace upon paper the magnified 
image of any microscopic object upon the microscope stage. 
It will be found of particular value in parasitology, wdiere it is 
required to study in detail the minute anatomy of organisms too 
large to be viewed at one time under the microscope. 

This apparatus is essentially a combination of mirrors and 
lenses by which the image of the sheet of paper is reduced by 
a suitable lens and is projected into the field of the microscope, 
so that the eye of the observer at the eye-piece of the camera 
lucida sees the image of the object and the paper and pencil of 
the examiner at the same time, i.e., apparently the enlarged 
object has been transferred to the sheet of paper, so that the 
pencil, as followed by the eye, can be made to trace the lines of 
the object upon the paper. 

The essential parts of the apparatus are the eye-piece, which 

2 



18 THE MICROSCOPE. 

is arranged to clamp firmly above the ocular of the microscope 
in the optical axis of the instrument. This contains mirrors and 
lenses suitably arranged to reduce and deflect the image pro- 
jected from the large reflecting mirror situated to the right of 
the ocular. This large mirror serves to reflect the image of the 
paper and pencil tip into the condensing lens of the camera 
lucida. 

In setting up this instrument, the circular clamp on the 
camera lucida is firmly attached to the upper part of the ocular, 
and the horizontal bar carrying the reflecting mirror is clamped 
into the guide provided for that purpose. The camera lucida 
is so hinged to the clamp that it can be swung out of the way 
while locating the proper field in the microscope, after which it 
can be swung into place and centered by means of two small, 
set screws. The reflecting mirror is also arranged so that its 
angle can be adjusted between forty-five and eighty degrees. 



II. 

THE SPUTUM. 



GENERAL CONSIDERATIONS. 

Sputum, or expectoration, is voided by coughing or clear- 
ing the throat. It is composed of the secretion and the exudate 
from the mucous membrane of the nose, pharynx, and trachea, 
down to the finest bronchioles and alveoli; also of material that 
may have entered the respiratory tract from adjacent organs 
(pus of abscesses and empyema) ; blood derived from anywhere 
along the respiratory tract ; and finally of material coming from 
the buccal cavity and from any part of the digestive tract. 

On account of this very complex origin the composition of 
the sputum is very variable. The sputum may be present, but 
may not be expectorated. Small children and occasionally 
adults, on account of bad habits, insufficient practice, or im- 
paired consciousness, swallow their sputum. In the majority 
of cases this difficulty can be remedied by teaching. For diag- 
nostic purposes the total output of sputum for twenty-four hours 
is collected in a suitable receptacle, and should be free from 
admixture of antiseptics. 

The sputum is examined in bulk with reference to its 
quantity, reaction, consistence, air-content, apparent composi- 
tion, color, and odor. 

THE AMOUNT. 

The amount of sputum voided in twenty-four hours may 
be very great. On the other hand, even when every effort is 
made to expectorate, very little is produced. Some phthisical 
patients, in spite of violent coughing, raise very little, and that 
is of a very tenacious quality. 

Scanty, or absent, sputum may be evidence of the first stage 
of bronchitis, asthma^ laryngitis or pleurisy. While in children 

(19) 



20 THE SPUTUM. 

under 6 or 7 years of age, it is usually absent because it is 
swallowed. 

Abundant sputum is of importance in a general way be- 
cause it denotes, in acute infectious conditions particularly, that 
nature is prompt to relieve the body of an abundant secretion, 
which, if retained, might cause serious consequences by further 
reducing the already diminished respiratory capacity. When 
large amounts of sputum are voided at short intervals, alter- 
nated with periods of practical absence, one may infer a tuber- 
culous, gangrenous or bronchiectatic activity, or rupture into 
the lung of an abscess of the lung, liver, kidney or subphrenic 
space. 

CONSISTENCE AND APPARENT COMPOSITION. 

The consistence of sputum may bear a certain relation to 
the amount when, if abundant, the consistence is lessened, and 
vice versa. This relation is by no means constant. 

Ordinarily, sputum is "slimy " It may, however, be serous, 
purulent, or bloody. The peculiar slimy characteristic of spu- 
tum depends upon the amount of mucus contained, while its 
stickiness depends, in part, upon the mucin and, in part, upon 
the proteid content. This is especially marked in lobar pneu- 
monia. 

(a) Watery, or serous, sputum, which is frequently blood- 
tinged, occurs in pulmonary edema, and in catarrhal influenza. 
Gastric disorders in neurotic old people may give rise to a thin, 
watery expectoration of considerable quantity, which is partly 
regurgitated and partly hawked up. 

(b) Viscid, or sticky, sputum, which adheres to the bottom 
of the container, even when completely inverted, is somewhat 
characteristic of lobar pneumonia, but may also be seen in 
phthisis, pertussis, and in broncho-pneumonia. 

(c) Mucous sputum, a clear diffluent sputum resembling 
egg-albumin, composed chiefly of mucus, is observed in the 
early stage of pneumonia, bronchitis, and phthisis; at the ter- 
mination of an asthmatic attack, in pertussis, pharyngitis, laryn- 
gitis, measles, and influenza. 

(cl) Muco-purulent sputum is composed of mucoid sputum 
in which occurs a varying number of streaks and masses of 



COLOB OF SPUTUM. 21 

opaque, yellow or greenish pus. It is noted toward the end 
of measles, pertussis, in resolving pneumonia, during phthisis, 
and in subacute and chronic bronchitis. 

(e) Purulent sputum, which is composed purely of pus, is 
rather rare, and, when observed, indicates rupture of a liver, 
kidney or subphrenic abscess, or purulent pleurisy into the 
respiratory tract. Opaque yellow sputum, consisting largely of 
pus, is found in bronchiectasis, phthisical cavities, broncho- 
pneumonia, and in the chronic or later stages of acute bron- 
chitis. 

(f) Numular sputum. Eing or coin-shaped masses of spu- 
tum, which sink immediately in water, occur at times in bron- 
chiectasis, chronic bronchitis or phthisical cavity. 

(g) Frothy sputum may be observed in bronchitis, bron- 
cho-pneumonia, and emphysema. Its most important relation 
is in pulmonary edema, in which condition it is full of air and 
resembles frothy soap-water. 

COLOR OF SPUTUM. 

(a) Rusty sputum, clue to evenly distributed altered blood, 
is generally indicative of lobar pneumonia, but may also be 
observed in tuberculosis pulmonalis. 

(b) Prune- juice expectoration, a rather fluid expectoration 
discolored by altered blood, is seen in gangrene and in cancer of 
the lung. 

(c) Current- jelly sputum is said to be characteristic of 
cancer of the lung. * 

(d) Black sputum. Sputum which is very dark or black 
streaked or specked, is found in persons who have inhaled coal 
dust or smoke for long periods of time. It is sometimes seen 
in gangrene of the lung. 

(e) Yelloiv, or green, sputum may be caused by abscess of 
the liver which has ruptured into a bronchus (bile pigment), 
and also in some cases of pneumonia (altered blood), and in 
pulmonary infections with chromogenic bacteria. 

(f) Shreds and casts may be observed in the sputum of 
chronic bronchitis, diphtheria, and rarely in pneumonia. Casts, 
unless large and branching (Fig. 4), are more apt to be found 



22 



THE SPUTUM. 



during microscopic search. Suspicious particles should be 
floated in water against a black background, and teased out 
with needles. 

(g) Blood-streaked sputum. Sputum streaked or discol- 
ored with blood, may be clue to violent vomiting or coughing, 
diseased tonsils or leakage from an aortic aneurism. Under 
these conditions it will appear as light red, but slightly altered 
blood. It may be present as a sequel of hemoptysis or abscess 




Fig. 4. — Bronchial Cast, from case of Fibrinous Bronchitis 
in service of dr. judson daland (original). 

of the lung in broncho-pneumonia, empyema or in bronchitis. 

If the blood is dark or black it may be due to pulmonary 
infarction. Most commonly hemoptysis is observed in phthisis, 
when it recurs intermittently for days or weeks. Finally, it 
should be remembered that malingerers may simulate disease of 
the respiratory tract by sucking blood from wounds of the gums, 
lips, tongue or cheeks. 

(h) Hemorrhagic sputum is observed in traumatic or 
tuberculous hemorrhage, in hemorrhagic infarctions, and in lobar 



AIK CONTENT, 23 

pneumonia. Also in tumors in or near the respiratory tract, 
and finally in congestion of the pulmonary circulation. 

Certain derivatives from the blood-pigment produce in the 
sputum shades very similar to that of blood — the rusty sputum 

of pneumonia, for instance. The coloring matter here is partly 
changed blood and partly a yellowish-red derivative of blood- 
pigment, about which very little is known. Peculiar lemon- 
colored and grass-green shades are also frequently observed in 
pneumonic sputum. These likewise are due to changed blood- 
pigment. Such sputa respond to Gmellin's test for bile-pigment 
(see page 214). A peculiar light-brown sputum is sometimes 
observed in heart disease, particularly mitral, in which amor- 
phous blood-pigment is found encapsulated in the alveolar epi- 
thelium (heart-failure cells). A type of green sputum has been 
observed in cases of lung tumor, the nature of the pigment in 
question is as yet unknown. 

(i) Extraneous discolorations. Other noticeable discolora- 
tions of the sputum are observed from the admixture of inhaled 
dust-particles. The black sputum of miners, and the blue spu- 
tum of workers in ultramarine, are examples of this class. 

Finally, it should be remembered that the sputum voided, 
following the ingestion of certain food-stuffs, may be discolored 
from contamination with these occurring in the mouth and 
pharynx. A greenish discoloration of the sputum is sometimes 
the result of the activity of certain chromogenic bacteria, espe- 
cially the Bacillus virescens. 1 Yellow and bluish sputa of prob- 
able bacterial origin have occasionally been observed. 

REACTION. 

The reaction of fresh sputum is generally alkaline; it may 
occasionally be acid, and usually becomes so after standing for 
some time through decomposition resulting from bacterial 
growth. 

AIR CONTENT. 

Sputum is often more or less foamy or frothy, due to the 
presence of air. Other things being equal, the content of air 
in the sputum is greater the finer the bronchi from which the 



^rick: Virchow's Archiv., Vol. cxvi., 1839. 



24 THE SPUTUM. 

sputum is derived. The consistence of sputum often has a bear- 
ing on this. The amount of air contained can easily be deter- 
mined by comparing its specific gravity to that of water. Air- 
containing sputum will float; airless sputum will sink. 

THE ODOR. 

Fresh sputum has a rather characteristic, but indescribable, 
odor. On standing it may acquire a disagreeably nauseating 
odor from decomposition, resulting from contained bacteria. 
Freshly voided sputum has a very decided odor in bronchiectasis, 
purulent bronchitis, tuberculosis, gangrene of the lung, and lung 
abscess. The disagreeable odor here arises from the activity of 
putrefactive bacteria. Stagnation of the secretion in cavities 
favors decomposition. In this way the foul odor of the expecto- 
ration in consumptives may be imparted to the breath. Finally, 
it must be remembered that fecal vomiting and diseased condi- 
tions of the mouth may be responsible for the odor of the breath 
and sputum, as will also the ingestion of certain volatile drugs,, 
including alcohol. 

HEMOPTYSIS. 

True hemoptysis means the expectoration of an appreciable 
amount of pure, or nearly pure, blood, not merely sputum tinged 
with blood. The amount of blood may continue small, and may 
persist for many days or may, as in the case of rupture of an 
aortic aneurism, be sufficiently large to cause death in a short 
time. 

Before making a diagnosis of hemoptysis it is necessary to 
exclude blood coming from the nose, pharynx, larynx, buccal 
cavity, and tonsils. 

The Common Causes of Hemoptysis. — (a) Pulmonary dis- 
ease, usually tuberculous. It may also occur in the early stages 
of lobar pneumonia, abscess, bronchiectasis, gangrene, and can- 
cer of the lung. 

(b) Cardiac disease. Here it may be the result of venous 
obstruction occurring in the course of valvular disease. This 
is a not uncommon cause of slight but long-continued bleeding. 

(c) Vascular disease. The most important condition is 



GROSS EXAMINATION OV SPUTUM. 25 

rupture of an aneurism into the respiratory tract. Leakage From 
the same cause may cause slight but persistent hemoptysis. 

(el) Discuses of (he blood. Hemoptysis may occur during 
the course of hemophilia, purpura, leukemia, scurvy, and severe 
anemia. It has been noted occasionally in the course of some 
of the exanthemata. 

(e) Miscellaneous causes. Vicarious menstruation and 
hysteria. 

GROSS EXAMINATION OF SPUTUM. 

Examination should be made both upon a white and a black 
background. Many sputa appear to the naked eye to be homo- 
geneous — pure mucus, pure pus, pure blood ; but sometimes not 
only may the sputum of one patient vary, but differences may 
be noted in each expectoration. 

"Dittrich's plugs' 7 are yellowish-white masses, the size of 
a mustard-seed, and are easily seen over a black background. 
They come from the smaller bronchioles in putrid diseases, 
especially in putrid bronchitis and pulmonary gangrene. Micro- 
scopically they are composed of clumps of bacteria and fatty 
acid, crystals. They have a very intense and disagreeable odor. 
Somewhat similar plugs may be encountered in the sputum in 
follicular tonsillitis. These plugs should not be confounded 
with those little masses described by, and known as, Cursch- 
mamr's spirals. (See page 27.) 

Formations consisting of fibrin are encountered in certain 
infections, and should be easily recognized by their white color,, 
tenacious consistence, and sometimes by their shape (casts and 
molds). 

Foreign bodies are sometimes aspirated into the respiratory 
tract, where they may remain for years without causing any 
symptoms until a fit of coughing dislodges them and they appear 
in the sputum. 

Concretions are sometimes, though very rarely, formed in 
the lungs during chronic inflammatory conditions. These are 
accidentally coughed up when they may be found in the sputum. 
Occasionally these stones may have their origin in the crypts 
of the tonsils, or they may be calcified lymph nodes that have 
ulcerated into the lung. 



26 THE SPUTUM. 

MICROSCOPIC EXAMINATION OF THE SPUTUM. 

A microscopic examination of the sputum reveals the pres- 
ence of cells, elastic fibers, casts, spirals, crystals, and micro- 
organisms. It is advisable to examine first the fresh unstained 
sputum, as the presence of fungi or crystals in the sputum, and 
the nature of many of the cellular elements, can be determined 
only in this way. Afterward dried and stained cover-glass 
preparations may be made for more minute and detailed study. 

Preliminary Examination. — This is necessary in order to 
locate suspicious particles which may be scattered throughout 
the large mass of sputum. This is made either with the unaided 
eye or with a hand-lens. A thin layer of sputum is necessary 
to successful examination. For this purpose a moderate-size 
Petri dish and cover is much better than the flat pieces of glass 
ordinarily employed, which are uncleanly and difficult to handle. 
A small amount of sputum is placed on the inside of the cover, 
and the other half of the dish jDressed down into this, the rim 
very successfully preventing the escape of excess of material. 

The Unstained Specimen. — For the purpose of isolating any 
characteristic particles, the sputum should be spread out in a 
thin layer in the dish, and the material teased out with needles 
or tooth-picks. Having located a likely particle, it is transferred 
to a clean slide and flattened out by pressing a cover-glass down 
upon it. This should be examined first by the low, and later by 
the medium, power objective. 

Appearance. — Most sputum consists, microscopically, of 
a ground-work of mucous matrix of indefinite structure and 
appearance, in which are imbedded a variety of microscopic 
objects, principally cells. 

1. Pus cells. The number of these indicates, in a general 
way, the purulent nature of the specimen. The character of the 
corpuscles varies greatly. Their size is from 7 to 10 micro- 
millimeters, they appear more or less granular, are sometimes 
distinctly pigmented, containing one or more irregular nuclei. 
The granules are composed some of proteid, some of fat, and 
some of extraneous debris. 

2. Epithelial cells found in the sputum differ from the pus 
cells, being usually larger in size, and by exhibiting one rather 



MICROSCOPIC EXAMINATION OF THE SPUTUM. 27 

large vesicular nucleus. Various types of epithelia arc met in 
the sputum, and their recognition is of considerable value in 
locating the origin of the expectoration, although, many times, 
the conditions to which they have been subjected after being shed 
have so altered their appearance that little knowledge can be 
gained from their study, (a) Squamous epithelia are derived 

1 rom the mouth, the pharynx, and from part of the larynx, (b) 
The cylindrical epithelium is derived from the nose or from the 
smaller bronchi, and are seen as pear-shaped or oval cells, some 
of which possess cilia, (c) Pulmonary, or alveolar epithelium, 
is oval and measures from 20 to 30 micromillimeters in diameter. 

3. "Heart-failure" cells. These are oval or round, pig- 
mented, alveolar cells. When numerous their presence is said 
to be indicative of chronic passive congestion of the lungs, usually 
depending on the failing compensation of cardiac valvular dis- 
ease. Their presence is, therefore, usually associated with the 
common signs in the lungs, which are indicative of failing com- 
pensation, viz. : moist rales, mucous expectoration, and cyanosis. 

4. Eosinopliiles may occasionally be found in large num- 
bers associated with Charcot-Leyden crystals in the expectora- 
tion of bronchial asthma. 

5. Red-blood cells. The appearance of red-blood cells in 
the sputum will depend largely upon the length of time that 
they have been shed. As they grow old they become pale, shad- 
owy, and fragmented. The finding of a few red-blood cells in 
the sputum is of no diagnostic import. They occur naturally 
in large numbers in hemoptysis, and are constant and more or 
less abundant in all inflammatory diseases of the lungs, par- 
ticularly phthisis. 

6. Casts. These may vary in size from those which repre- 
sent molds of the trachea and larger bronchi to those coming 
from the smaller bronchioles, and which are from 14 to 1%' 
inches long. These smaller casts are the more common, and 
when present usually require the use of the low-power -objective 
to demonstrate them. These casts usually occur in one of the 
three f ollowing diseases : the largest in diphtheria, medium- 
size in fibrinous bronchitis, and the smallest in lobar pneumonia. 

7. Curs ch ma nil's spirals consist of worm-like spirals 1 to 

2 centimeters long, and about 1 millimeter wide. They are 



28 THE SPUTUM. 

more or less opaque, and are usually found surrounded by a 
thick, clear mass of mucus. They frequently show a central, 
undulating, thread-like core around which are twisted, in a 
spiral manner, the mucous threads. Entangled in these spirals 
are usually eosinophiles and Charcot-Leyden crystals. They 
occur frequently in the sputum of bronchial asthma, more rarely 
in phthisis, bronchitis, and in lobar pneumonia. Their presence 
may be of service in differentiating bronchial from other forms 
of asthma. 

Crystals. — (a) These are found usually only when the 
sputum has been retained within the body for a length of time. 
Crystals of fat or of fatty acids are most frequently encoun- 
tered. They appear as long, slender needles, either singly or 
grouped into fine rosettes or sheaves. They are readily soluble 
in potassium hydrate or in ether. This solubility is easily de- 
termined by allowing a little of either fluid to flow under the 
edge of the cover-glass while observing the crystals in question. 

(b) Crystals of calcium phosphate may be encountered 
under conditions of retention and stagnation. 

(c) Charcot-Leyden crystals are occasionally encountered, 
particularly in the expectoration of bronchial asthma, and are 
here accompanied by eosinophiles. They appear as colorless, 
elongated double pyramids, varying considerably in size. They 
are often so small that high magnification is required to reveal 
them. 

(d) Choi est erin crystals are but rarely seen in the sputum. 
They occur as transparent, colorless, rhomboiclal platelets, with 
notched or irregular angles and ends. 

(e) Hemato'idin crystals are derived from hemoglobin by 
a process of decomposition, and occur as needles and rhomboidal 
platelets of reddish and brownish hue. They are found chiefly 
in the sputum from old abscesses or perforating empyemas. 

(f) Leucin globules and ty rosin crystals are found in 
putrid sputum from old perforating abscesses or in putrid bron- 
chitis. 

(g) Calcium oxalate, in minute octahedral crystals, are 
occasionally met. 

Elastic Fibers. — When the lung-tissue is destroyed to 
any extent by pathologic processes, elastic fibers are apt to be 



PREPARATION OF THE STAINED SPECIMEN. 9 ( .) 

encountered in the sputum. Their presence in the sputum 
proves conclusively the occurrence of some destructive process 

within the lung. Hence their importance in the diagnosis of 
tuberculosis of the lung before the appearance of tubercle ba- 
cilli. Elastic libers occur also in pulmonary abscess and gan- 
grene. 

These fibers are usually detected in a thin layer of sputum 
examined microscopically. In this examination care must be 
observed to avoid confusing true elastic fibers with the some- 
what similar vegetable fibers, which latter are generally larger 
and less uniformly wavy. 

To detect particles or shreds of elastic tissue in the sputum, 
suspicious lumps should be thoroughly mixed with an equal 
quantity of 20-per-cent. sodium hydrate solution; then a large 
volume of water is added, and the whole allowed to sediment 
for a few hours. The sediment is then removed and examined 
under the microscope for the characteristic fibrillated masses. 

If elastic tissue is not found by this procedure, the entire 
quantity of the twenty-four-hour specimen should be boiled 
with an equal quantity of the sodium hydrate solution. The 
resulting gelatinous mass is then mixed with several volumes 
of water, and allowed to sediment. About 15 cubic centimeters 
of the sediment is now removed with a pipette and centrifuged 
for fifteen minutes. The final precipitate is now carefully 
removed and examined as above. 

Fragments of toiors are occasionally encountered in the 
examination of sputum. These should be removed and pre- 
pared for sectioning and staining. 

PREPARATION OF THE STAINED SPECIMEN. 

Another suspicious particle having been isolated and re- 
moved from the mass of sputum, it is transferred to and care- 
fully spread upon a clean cover-glass. This should then be 
treated to fixation and staining, the technic of which will de- 
pend upon the nature of the information sought. (For meth- 
ods of staining and for differentiation of the organisms, see 
section devoted to Bacteriologic Methods,' page 244.) 



30 



THE SPUTUM. 



PULMONARY ACTINOMYCOSIS. 

This condition, also known as disease of the ray-fungus, 
occasionally causes disease of the lung, but is exceedingly rare 
in this country. The characteristic yellowish- or grayish-green 
granules, if found, are often sufficient for a diagnosis which 
should, however, always be confirmed by microscopic search. In 
some cases the characteristic microscopic rosettes with clubbed 
rays are found; in others only branching threads, staining by 
Gram's method, will be found. 

Clinically, the course of the disease is similar to that of 
pulmonary tuberculosis, except that instead of the tubercle 
bacillus, the ray-fungus is found. 

ECHINOCOCCUS. 

Barely echinococcus booklets enter the pulmonary tract and 
appear in the sputum. They usually originate in abscesses of 
adjacent organs, particularly the liver. (See section, "Animal 
Parasites," page 101.) 

DISTOMUM PULMONALE. 

This is a disease of the lungs, usually occurring in Asia, 
but occasionally encountered in other parts of the world. It is 
due to the presence of a worm which, by causing hemorrhage, 
may be mistaken for phthisis. When present its eggs can always 
be demonstrated in the sputum. They are oval, of a brownish- 
red color, 0.08 millimeter long, and 0.056 millimeter broad, sur- 
rounded by a cuticle. (See section, "Animal Parasites/' page 
101.) 



III. 

THE BLOOD. 



THE CHEMICAL COMPOSITION OF THE BLOOD. 

A general idea of the composition of the blood may be 
had from the following table, which is taken from Simon's 
"Physiologic Chemistry/ 5 The calculations are made for 1000 
parts by weight. 

Corpuscles 480. 00 parts 

Water 276.90 » 

Oxyhemoglobin 193.90 » 

Stroma, including salts 9.20 " 

Plasma 520.00 parts 

Water 477.37 " 

Albumins 35.88 " 

Extractives 2.39 " 

Inorganic salts 4.36 " 

The predominating solid substance in the blood is oxy- 
hemoglobin; it represents 10 per cent, of the total weight of 
the blood, 10 per cent, of the weight of the corpuscles, and 65 
per cent, of all organic matter present. 

The mineral constituents comprise sodium, potassium, cal- 
cium, magnesium, and iron. 

Fats are present to the extent of from 0.2 to 0.3 per cent. 
These may be temporarily increased after the ingestion of much 
fatty food, and also in many pathologic conditions. . 

The plasma normally contains small amounts of oxygen 
and nitrogen in solution, with varying amounts of carbon 
dioxide. 

METHODS OF OBTAINING BLOOD FOR EXAMINATION. 

The tip of the finger or lobe of the ear are the sites usually 
selected from which to obtain specimens. In the majority of 

(31) 



32 



THE BLOOD. 



examinations only a small amount — a few drops — is necessary. 
This is obtained by simple puncture of the skin made with a 
glover's needle, the half-point of a new steel pen, . a Dal and 
lancet (see Fig. 5), or the so-called pistol-knife. The two 
last-mentioned instruments are to be preferred because they 
permit of regulation of the depth of the puncture. 

For larger quantities of blood, a few cubic centimeters or 
more, it is advisable, provided there is no contraindication, to 
obtain the specimen by the use of wet cups. It must be borne 




Fig. 5.— Fleischl Hemoglobinometer, surrounded by accessories 

necessary to performance of the test, 

including daland lancet. 



in mind that by this method there is always more or less ad- 
mixture of lymph. Another method is to draw the blood from 
a dilated vein into- a large sterile antitoxin syringe. Lastly, an 
ordinary venesection may be resorted to. 

The withdrawal of blood, if aseptically performed, is prac- 
tically free from danger and need disturb the patient very little. 
Before proceeding in any case it is advisable to determine the 
absence of hemophilia in the patient. 

Appearance of Fresh Blood. — The exuding drop of blood 



THE SPECIFIC GRAVITY. 33 

shows even to the naked eve a number of properties. The redder 

it is. the richer it is in oxyhemoglobin; the darker, the greater 
the amount of reduced hemoglobin. Microscopically, it reveals 

a great number of cellular elements; these are colored and col- 
orless discs. 

The red cells appear as non-nucleated bi-concave discs, 
measuring on the average 7 micro-millimeters in diameter. 
Viewed singly through the microscope by transmitted light, they 
are of pale-greenish hue. The colorless cells or white corpus- 
cles are, as a rule, somewhat larger than the red cells, and pre- 
sent either mono- or poly-nucleated protoplasm. 

The plaques, or blood-platelets, appear as minute, color- 
less discs measuring less than half the diameter of the red cells. 
They usually occur in groups or bunches of half a dozen or 
more, and are present in normal blood to the number of about 
635,000 per cubic millimeter. 

There are no other morphologic constituents of the blood. 

Color. — The color of normal blood is due to the presence 
of an albuminous substance in the corpuscles termed hemo- 
globin. In the arterial blood it is in combination with oxygen, 
and is here termed oxy-hemoglobin. In the venous blood a mix- 
ture of both hemoglobin and oxy-hemoglobin occurs. With a 
preponderance of oxy-hemoglobin, the blood tends to a scarlet 
hue ; when the hemoglobin predominates, the blood is of a 
bluish color. 

Pathologic Changes in Color. — In coal-gas poisoning the blood is cherry- 
red. After poisoning from potassium chlorate, aniline, hydrocyanic acid, 
and nitro-benzol, the blood is brownish-red or chocolate color. In extreme 
cases of leukemia the blood may have a milky appearance due to the 
excessive number of white blood cells present. 

The Odor. — This is characteristic and differs in different 
species of animals. It is due chiefly to the presence of volatile 
fatty acids. 

The Taste. — The taste of blood is salty, but at the same 
time insipid. 

THE SPECIFIC GRAVITY. 

The specific gravity seems to vary with the amount of 
hemoglobin. It is influenced by the age and sex of the indi- 



34 THE BLOOD. 

vidual, the process of digestion, exercise, pregnancy, etc. The 
normal average in adults varies between 1.058 and 1.062. 

Determination of the Specific Gravity (method of Ham- 
merschlag). — A cylinder about 10 centimeters in height is 
partly filled with a mixture of benzol (sp. gr. 0.889) and chlo- 
roform (sp. gr. 1.526), so that the specific gravity of the mix- 
ture lies between 1.050 and 1.060. Into this a drop of blood is 
allowed to fall directly from the finger. It is then brought into 
suspension by the addition of either a little chloroform or benzol, 
according to the tendency of the drop to sink or rise in the 
cylinder. 

As soon as the drop remains stationary in the fluid the 
specific gravity of this is taken by an accurate hydrometer (one 
reading to the fourth decimal should be used). The reading 
represents the specific gravity of the blood tested. 



THE AMOUNT. 

The total amount of blood in the normal adult is said to 
amount to about one-twelfth or one-fourteenth of the body 
weight. 

THE REACTION. 

The reaction of the blood is slightly ' alkaline, due to the 
presence of the mono-sodium carbonate and the di-sodium 
phosphate in solution in it. The reaction may be roughly 
determined by drawing a strip of neutral litmus paper, which 
has been thoroughly moistened with a concentrated solution of 
common salt, through the blood, and then rapidly washing the 
corpuscles off with the same solution. Owing to the develop- 
ment of certain acids, the alkalinity of the blood rapidly dimin- 
ishes after it is shed. This fact renders this determination a 
rather difficult matter. The normal variation of alkalinity is 
very slight. By accurate titration the normal degree of alka- 
linity of the blood, under normal conditions, corresponds to 
from 325 to 360 milligrams of sodium hydrate for every 100 
cubic centimeters of blood. 



ESTIMATION OF HEMOGLOBIN. 35 

QUANTITATIVE CLINICAL METHODS. 

The usual clinical methods applied to the blood in the 
study of disease are: the estimation of the percentage of 
hemoglobin, the enumeration of the erythrocytes and the white 
blood-corpuscles, and a differential count of the various white 
elements. 

The apparatus necessary for the performance of these sev- 
eral examinations are as follows: — 

I. — A good microscope with 3, 6, and oil-immersion (Y^) 
objectives. 

• II. — A hemoglobinometer (Gower's, FleischPs or Sahibs). 

III. — A Thoma-Zeiss hemocytometer. 

TV. — Slides, covers, stains, etc. (For complete list see 
Appendix.) 

Estimation of the Percentage of Hemoglobin. — Method of 
Gower: The tip of the finger or lobe of the ear is punctured 
after having been thoroughly cleansed with alcohol, followed by 
ether. The first drop obtained is wiped away, and the second, 
which should flow without assistance either by pressure or rub- 
bing, is drawn up by suction into the pipette to the 20 centi- 
meter mark; all adhering blood is wiped from the outside of 
the tube. The contents are now immediately forced out into 
the graduated observation tube, which has previously had a 
few drops of distilled water placed in it (this is to prevent the 
blood from coagulating on its walls). Be sure that all blood 
contained in the mixing tube has been washed into the gradu- 
ated tube. K~ow, while holding the two tubes side by side 
directly against the light, acid distilled water, drop by drop, 
until the shade of color is the same in the two tubes. The 
division on the scale to which the fluid rises will express the 
per cent, of hemoglobin. 

Method of Fleisch. — The instrument (Fig. 5) consists 
of a metal stand having a horizontal table with a circular aper- 
ture in its center, beneath which is placed a reflector of plaster- 
of-Paris. Immediately beneath the aperture and above the 
reflector, a graduated wedge of tinted glass is arranged to move 
in a horizontal plane by means of a milled thumb-screw. This 
graduated wedge of glass is shaded to represent varying degrees 



36 THE BLOOD. 

of hemoglobin content, and carries with it a scale indicating 
the blood-strengths. For collecting and diluting the blood for 
examination, a pipette and diluting chamber are furnished. 
The teclinic is as follows: — 

By the aid of the pipette which is furnished with the ap- 
paratus, an exactly determined amount of blood is dissolved in 
a measured quantity of distilled water (the contents of one-half 
of the diluting chamber, the other half being filled with an 
equal amount of distilled water). The cell thus prepared is 
then placed over the aperture in the table, and artificial (candle, 
lamp or gas) light is directed through it by means of the re- 
flector. The part of the wedge is now searched for, which 
accurately compares in tint with the solution of blood under 




Fig. 6.— Thoma-Zeiss Hemocytometer in Case. (A. H. Thomas.) 

examination, and the number of the scale is read off that corre- 
sponds to this point on the glass wedge. It is a matter of com- 
mon experience that the grading of these instruments is too 
high, and that a specimen of blood that corresponds to 90 or 
95 on the scale is normal. 

Enumeration of the Corpuscles. — The Thoma-Zeiss ap- 
paratus consists of a counting slide and two diluting pipettes, 
termed "melangeurs" (Fig. 6). This slide is so constructed that 
it contains a chamber in its center having a depth of %o milli- 
meter. The center of the floor of the cell is divided by fine 
microscopic lines into minute squares, the sides of which are 
equal to % millimeter. The cell is completed by the applica- 
tion of a special cover-glass. Under these conditions each cube 
as outlined has a capacity of /4ooo cubic millimeters. 



ESTIMATION OF ERYTHROCYTES. ;>; 

Those small squares are divided into groups of sixteen by 

means of double ruled lines, each group of sixteen small squares 
constituting a large 1 square. 'There are sixteen of these large 
squares in the slide. There are, therefore, two hundred and 
fifty-six small squares in the sixteen large squares. If we in- 
clude also the small squares forming the boundaries between 
the large squares, the total number of small squares will be 
four hundred (Fig. 9). This counting chamber is used in esti- 
mating both the red and the white cells, also the number of 
cells in the cerebrospinal fluid, pleural effusions, etc. 

Estimation of the Erythrocytes. — The "melangeur" having 
the smallest bore and having marks at 0.5, 1.0, and 101, is 
used in this estimation. The drop of blood issuing from the 
puncture is drawn up to the 0.5 mark, the tip then quickly 
freed from adherent blood, and immersed in a 2%-per-cent. 
solution of potassium bichromate (or Hayem^s solution, for 
which see Appendix), which is drawn up to the 101 mark. 
The tip of the tube is now stopped with the finger and the tube 
vigorously shaken, for at least a minute, to insure thorough 
and even dilution of the blood. The portion of the diluting 
fluid contained in the capillary part of the tube is blown out 
and wiped away, and the next drop of the mixture placed in the 
center of the floor of the counting chamber. This drop should 
be entirely free from bubbles. The cover-glass is now applied 
over this and pressed down firmly around the edges until Xew- 
ton's rings appear. 1 A few r moments should now pass before 
counting to allow the corpuscles to settle to the bottom of the 
chamber. 

A simple and practical method of arriving at the: number 
of red corpuscles obtained in a cubic millimeter of blood is the 
following : Select and count from the various parts of the cham- 
ber five large squares (Fig. 7), counting the border cells only 
upon- the upper and right-hand lines. The total of small 
squares counted will be eighty. To the total of the corpuscles 
counted in the five large squares add four 0000, and the number 
resulting w T ill be the number of red corpuscles in 1 cubic milli- 
meter of undiluted blood. 



] A series of curved, colored lines appearing behind the plane glass surfaces, 
where they are firmly pressed together. 



38 



THE BLOOD. 



Enumeration of the Red Cells. — Explanatory note : y± 000 cc. 
equals the cubic capacity of one small square. y 200 equals the dilution of 
the specimen of blood. 'Five large or eighty small squares is the number 
of squares counted. Then the number of cells per cubic millimeter in the 
undiluted specimen will equal the number of cells in one small square 
multiplied by the dilution times 4000, viz: — Let x equal cells per cubic 
millimeter, and let y equal number of red blood- cells in eighty small 
squares. Then, ■/-$ X 200 X 400 = x which simplified is the same as 
y X 10000 = x = the number of red blood -cells in one cubic millimeter 
of undiluted blood. 

A Second Method. — Count all the small squares (-±00) ; 
divide the number of cells counted by the number of squares. 
This will give the average number of cells in one square ( /4ooo 
cubic millimeter) of diluted blood. To determine the number 




Fig. 7.— Appearance of Field of Thoma-Zeiss Hemocytometer when 
properly mounted for counting the red corpuscles. 



of cells in 1 cubic millimeter of undiluted blood, it is only 
necessary to multiply this by -1000 and this product by the dilu- 
tion. 

Example. — Suppose 1200 cells have been counted in 400 
squares, then the average in one small square is three; this, 
multiplied by the number 4000 will equal 12,000, the number 
of corpuscles contained in 1 cubic millimeter of diluted blood, 
since the dilution was 200 times; then 12,000 x 200 will equal 
2,400,000 red blood-cells in 1 cubic millimeter of undiluted 
blood. 

Eecently Drs. Einhorn and Laporte have perfected a rapid 
method of counting by means of a specially constructed count- 
ing diaphragm. 



THE HEMATOKRIT. 39 

The normal number of erythrocytes is usually placed at 
5,000,000, although it is not uncommon to find the normal in 
many patients maintained above this figure. 

Determination of the Erythrocytes by the Centrifuge. — 
The Daland hematokrit (Fig. 8) offers a quick, simple, and 
accurate method of determining the number of red corpuscles 
in the blood. The two great advantages of this process are, 
first the elimination of the personal equation, and second, as 
there is no dilution of the blood required, the frequent source 
of error arising from this cause does not enter in. 

The hematokrit consists of an extremely light, though very 
strong, metal frame containing two glass tubes, each 50 milli- 
meters long and graduated in one hundred parts, and having 
a uniform lumen of 0.5 millimeter. 

The length of the percentage tubes and their distance from 
the center of the revolving hematokrit frame must always be 




Fig. 8.— Daland Hematokrit, showing one percentage 
tube in position. (a. h. thomas.) 

the same as was used in the original experiments by which the 
comparative determinations were made, since any variations in 
these factors will produce corresponding variations in the 
results obtained. 

The Techxic. — The blood having been secured as out- 
lined on page 31, the blunt end of the graduated percentage 
tube is attached to a suction pipette and the tube completely 
filled with freshly drawn blood. After any excess of blood has 
been carefully wiped away, the blunt end of the tube is placed 
in the distal cap of the hematokrit frame, and the other end 
pressed down upon the inclined plane of the proximal cap until 
it falls into and locks in the cavity. The second tube should 
be similarly treated and placed in the opposite side to serve as 
a check. It is absolutely necessary that the entire procedure 
should be performed with celerity so as to anticipate coagula- 
tion. After filling, immediately place the frame upon the spin- 
dle of the high speed centrifuge, and turn the handle at the uni- 



40 THE BLOOD. 

form rate of eighty turns per minute for exactly two minutes. 
This will produce 10,000 revolutions per minute in the spindle. 
At the expiration of this time all the corpuscles will be found 
to occupy the distal end of the tube, a thin, almost invisible, 
whitish line of white corpuscles appears between them and the 
plasma, which will be found to occupy the proximal portion of 
the tube. With the aid of a hand lens the height of the column 
of red corpuscles is read off, and the reading multiplied by 100,- 
000, which converts the volume percentage into the number of 
corpuscles per cubic millimeter of the blood under examination. 

The speed of the handle of the centrifuge must be uniform 
throughout the whole period, because this rate of rotation is 
the measure of a definite amount of centrifugal force gener- 
ated in the revolving arms of the hematokrit frame. If the 
rate of rotation is either increased or decreased, the height of 
the column of cells will be similarly affected, thereby producing 
either too low or too high a reading. 

A possible source of uncontrollable error is the variation 
in size of the individual erythrocytes which occurs in certain 
pathologic states. The most accurate results by this method 
will therefore be obtained when the corpuscles are of uniform 
size. This fact has been taken advantage of in the estimation 
of the volumetric quotient, and its relation to diagnosis and 
prognosis in certain blood-conditions (see page 41). 

The Percentage of Erythrocytes. — To obtain the percent- 
age of red blood-corpuscles, take the first two left-hand figures 
of the count in 1 cubic millimeter and multiply these by two ; 
this will represent, approximately, the percentage of red blood- 
cells. 

Explaxatiox. — If we consider normal blood to contain 
exactly 5,000,000 corpuscles, then the figure 50 x 2 would equal 
100 per cent., or normal. If, however, only 4,790,000 were 
counted in 1 cubic millimeter of blood, then 47 x 2 will equal 
94 per cent, of erythrocytes. 

The Color-Index. — From a knowledge of the percentage 
of hemoglobin and the percentage of cells, we are in a position 
to calculate the average color-index per cell, i.e., its relative 
hemoglobin-content as compared with the normal cell. 

Example. — If the percentage of hemoglobin be 100 and 



THE VOLUME-INDEX OF ERYTHROCYTES. 41 

the percentage of cells 100, then the color-index of each cell 

would be 1.0 or normal, viz. : — 



loo ( ( hemoglobin 
100 <y c red cells " 



: color-index 1.0. 



If the percentage of hemoglobin be 67, and the percentage 
o( corpuscles 90, then each individual cell will contain less than 
a normal amount of hemoglobin: — 



^& L 



67 % hemoglobin 



90 f c red cells 



= color-index 0.74. 



The Volumetric Quotient or the Volume-Index of the 
Erythrocytes. — The investigations of Capps 2 have demonstrated 
that the normal volume obtained by the centrifuge hematokrit 
is 50 per cent, in the normal specimen. This volume he desig- 
nates as 1. The volume in pathologic alterations can be cal- 
culated in percentage of the normal volume, just as in the 
amount of hemoglobin estimated under similar circumstances. 
Having determined the volume by the hematokrit the erythro- 
cytes are next counted and their number expressed in percent- 
age by comparing with the normal. By dividing the volume of 
erythrocytes by the number of erythrocytes in the same blood 
(both expressed in percentage), w r e obtain the "volume-index" or 
"volume-value" of the erythrocytes, which is analogous to the 
color-index or hemoglobin as obtained above. This number in a 
measure of the average volume of the individual erythrocyte, 
and obviously under normal conditions, equals 1. Capps found 
that an increase in the volume-index of the erythrocytes is one 
of the most constant and accurate characteristics of pernicious 
anemia, which agrees w^ith the well-known fact that many 
macrocytes appear in the blood in this disease, and that the 
color-index is also greater than one. In contrast to this, the 
so-called secondary enemas usually show a diminished volume- 
index ; the same is true of chlorosis, in which affection the 
volume-index may be taken into account in determining the 
prognosis, since a normal or only slightly decreased volume- 
index gives a favorable prognosis, whereas a markedly dimin- 
ished volume-index may be considered a bad prognostic sign. 



2 Jour. Med. Research, Vol. x 3, Boston, 1903. 



42 



THE BLOOD. 



When utilized in this way the volume-index may become 
a more reliable sign in chlorosis than is the percentage of hemo- 
globin or the color-index 3 

Estimation of the Leukocytes.— The Thoma-Zeiss appar- 
atus is used for this estimation. The "white" pipette is of 
larger calibre than the "red" pipette, and makes a lower dilu- 
tion (1 to 10 or 1 to 20). For diluting it is customary to use 
a 0.3- or 0.5-per-cent. solution of acetic acid. This solution 
preserves the white cells, at the same time decolorizing the 
erythrocytes, thus facilitating enumeration. 

Tor mixing and preparing the slide the same method is 




Fig. 9.-Ruling of Chamber of Thoma-Zeiss Hemocytometer. 

employed as described for the red cells. It is customary to 
count the total number of squares (400) (Fig. 9), and to mul- 
tiply the total number of cells counted by two hundred; the 
result will be the total number of white cells per cubic milli- 
meter of blood. 

Enumeration of the Leukocytes. — Explanatory note: %ooo ce - 
equals the cubic capacity of one small square. y 20 equals the dilution of 
the blood, and 400 (all) the small squares counted. Then the number 
of leukocytes per cubic millimeter of undiluted blood will be found by 
dividing the number of cells counted by the number of squares counted 
and multiplying by the dilution times 4000 viz:— Let x equal the number 
of leukocytes per" cubic millimeter, and y equal the number of cells 
counted in 400 small squares. Then, jjj X 20 X 4000 = x or, y X 
200 = x = leukocytes per cubic millimeter of undiluted blood. 

3 Sahli's Diagnosis quoting: Capps; loc cit. 



QUANTITATIVE CLINICAL METHODS. 40 

Examination of a Drop of Fresh Blood. — If a drop of fresh 

blood is placed upon a slide and a perfectly clean cover-glass 
allowed to fall upon this, a fresh preparation for examination 
will be produced, which may either he examined immediately 
or it' sealed around the edge with a little vaseline, may be car- 
ried to the laboratory and examined within an hour. With the 
aid of a 6 objective many interesting points may be observed. 
First, the shape of the red cells and their hemoglobin staining, 
also rouleaux formation. Second, the structure of the white 
corpuscles by which the different varieties may be roughly dif- 
ferentiated. Third, the blood-platelets. Fourth, a rough idea 
of the relative number of red and white cells may be formed. 
This matter will be discussed more in detail a little later. 

Studies of the minute structure of the leukocytes cannot 
be made satisfactorily by this simple method, as some of the 
forms are present only in small numbers, and are detected 
with difficulty. Further, prolonged examination of the fresh 
specimen allows time for changes, which rapidly obscure the 
identity of the cells ; hence we are obliged to resort to a method 
of preparing a specimen which shall be permanent and which 
can be stained as a further aid to differentiation. These meth- 
ods have the added advantage that they allow the investigator 
to work quite independently of the presence of the patient, to 
choose the time and place of examination, and permit at any 
time of verification and demonstration of the original result 
obtained. 

Preparation of the Specimex. — The blood is obtained 
from the tip of the finger or the lobe of the ear after the man- 
ner described above. Cover-glasses should previously be pre- 
pared for reception of the fresh blood as soon as it is drawn. 
The cover-glasses should be square, of seven-eighths or one inch 
in diameter, and of special thinness (0.1 to 0.08 mm.). They 
should be washed carefully with warm water and soap, dried 
with fat-free gauze, and finally wiped off with ether. This is 
necessary to insure proper spreading of the blood between the 
two glass surfaces, which would be prevented by traces of fat 
or particles of fiber or dust, etc. Just before the glasses are 
used they should be wiped off with a piece of silk or tissue 
paper, and thereafter handled with care by the corners, or, 



44 THE BLOOD. 

better still, with forceps. This preparation of the cover-glasses 
may be done in the laboratory or carried on at the bedside, as 
desired. 

The glasses being ready and the finger cleansed and punc- 
tured, a small drop of blood is allowed to exude without pres- 
sure. Now a cover-glass is taken up diagonally between the 
thumb and finger of one hand, and its center allowed to touch 
the drop of blood. Immediately taking another cover-glass in 
the same manner in the other hand, it is allowed to fall upon 
the drop. If the glasses are perfectly clean and the maneuver 
properly executed, the blood will immediately spread out in a 
uniform layer between the two glasses. The two cover-glasses 
are now separated one from the other by a rapid sliding motion, 
in the strict plane of their surfaces, without the slightest lifting 
or tilting. 

If the drop taken was small enough and the technic prop- 
erly carried out in every detail, the result will be two uniform 
smears, covering the larger portion of the surfaces of the cover- 
glasses which, examined under the microscope, will present a 
uniform layer of corpuscles, with little if any overlapping of 
the individual cells. 

Usually six films are prepared in this manner and allowed 
to dry in the air without heat. 

Methods of Fixatiox. — In using the Eomanowski stains 
no previous fixation of the albuminous film is required. Before 
applying any of the other stains it is necessary to "fix" the 
blood-film. If this is not done the corpuscles will be washed 
from the glass by the fluids applied. 

1. A strip of sheet copper eighteen inches long and two 
or three inches wide, is placed upon a stand, and the flame from 
a Bunsen burner placed under one end. After allowing suf- 
ficient time for the plate to attain a maximum heat the films 
are placed upon it at a point where a drop of cold water fails 
to roll off but adheres to the hot metal and steams away. At 
this point fixation is complete in from one-half to three-quarters 
of an hour. 

2. Fixation may be accomplished by immersion for five to 
ten minutes in a mixture of 1 part formaldehyde solution and 
99 parts absolute alcohol. 



STAINING METHODS. 45 

o. For routine work when fair accuracy, accompanied by 
speed in obtaining results, is necessary, fixation in the naked 
flame of a Bunsen burner may be practiced. This is accom- 
plished by holding the dried smear diagonally between the thumb 
and forefinger of one hand, film side up, and passing it through 
the flame a number (five to ten) of times with sufficient rapid- 
ity to prevent the fingers being burned. 

Methods of Staining: Eosin and Methylene-Blue. — The 
fixed film held in the cover-glass forceps is flooded with a 
1 o-per-cent. alcoholic solution of eosin, which is gently washed 
off with distilled water after the lapse of one or two minutes. 
The counter-stain of methylene-blue, 1-per-cent. aqueous solu- 
tion, is then added and allowed to act for from two to five 
minutes,, according to the density of the film. Finally, wash- 
ing in distilled water, blotting and drying, complete the process 
prior to mounting and examination. 

This is an extremely simple, yet effective and permanent, 
method of staining blood-smears. Little difficulty will be ex- 
perienced in obtaining satisfactory specimens. The eosin should 
not be allowed to act too long, but there is little danger of over- 
staining with the methylene-blue. Both of these stains are 
permanent when prepared, and can be kept until exhausted. 

Properly performed this method gives very vivid and con- 
trasting pictures, the nuclei of the different white cells tak- 
ing various shades of blue, pale in the lymphocytes and dark 
in the polymorphonuclears. The protoplasm appears faintly 
tinted blue (paler than the nuclei). The eosinophilic granules 
are a bright pink. The erythrocytes a dusky red. 

EhrlicVs Triacid Staining Method (for formula and pupa- 
tion see Appendix). — The smear is taken in the cover-glass 
forceps and a few drops of the prepared stain placed upon it 
and allowed to remain for about five minutes. There is little 
danger of over-staining. The specimen is next washed in dis- 
tilled water and thoroughly dried prior to mounting. If 
mounted in Canada balsam it is imperative that this should 
not contain any chloroform, otherwise the colors will gradually 
become blurred. 

The Romanowski or "Universal" Staining Method. — Leish- 
mann's modification of the Eomanowski stain, as made by 



46 THE BLOOD. 

Wright, 4 is the stain used. This method is altogether quicker 
and easier than the method of Ehrlich. It requires no fixing 
fluid or heating apparatus, and gives pictures which are uni- 
formly superior. 

Technic. — Allow three or four drops of the prepared stain 
to fall upon the smear and permit it to remain one-half min- 
ute, rocking the cover gently so as to insure an even distribu- 
tion of the stain. Xo attempt is made to check evaporation. 
At the end of one-half minute add double the quantity of dis- 
tilled water, i.e., six to eight drops, and allow it to mix with the 
alcoholic stain. Immediate mixing is hastened by gently rock- 
ing the cover-glass. The film is now allowed to stain for five 
minutes. In thick smears ten minutes may be necessary. The 
stain is now gently washed off with distilled water, and a few 
drops of water are allowed to rest on the film for one minute, 
when it should be dried in air and mounted. 

Appearance of Blood-Films. 5 — Erythrocytes, pale pink 
or greenish semi-transparent. 

Polymorphonuclears. — Xuclear network, stained a very 
ruby-red color, or with sharply defined margins. Extra-nuclear 
protoplasm, colorless. Fine eosinophilic granules, red. 

Mononuclears. — Xuclei ruby-red with extremely sharp, 
clear outlines. Extra-nuclear protoplasm, pale eau-de-nid or 
blue, occasionally showing a few red granules. 

Lymphocytes. — Same as mononuclears, except that as a 
rule the nuclei are more deeply stained. 

Coarse-Grained Eosinophils. — Xucleus only, red, but not 
so densely stained. Granules pale pink. 

Basophil es. — Granules very densely stained of a very pur- 
plish-black tint. Xucleus red, but usually more or less meshed 
by granules overlaying it, 

Nucleated Bed Cells. — Xucleus almost black, with sharp 
outline, extra nuclear portion gray. 

Blood Plates. — Deep ruby-red with spiky margins, fre- 
quently showing a pale blue spherical zone surrounding the red 
center. 

Bacilli and Micrococci. — Speaking generally, these stain 

4 Wright: Jour. Med. Research, Vol. vii, 1902. 

5 British Medical Journal. Vol. ii, page 757, 1901. 



TERMS IX COMMON USE IX CLINICAL BEMATOLOGY. 47 

evenly blue, but by prolonging the staining period and subse- 
quently decolorizing with absolute alcohol, many interesting 
variations may be noted with different organisms by which 

structural details are brought out not generally observed by 
other staining methods. 

Malarial Parasites, — The body of the parasite stains blue 
and its chromatin ruby-red. In the case of the tertian parasite, 
Schuttner's dots are well marked in the containing red blood- 
corpuscles. 

The only weak points in the Komanowski stain are said 
to be the deceptive resemblance between megaloblasts, certain 
lymphocytes and certain myelocytes, and failure to differentiate 
the basophiles. 

TERMS IN COMMON USE IN CLINICAL HEMATOLOGY- 

Anemia. — A condition of the blood in which there is a 
deficiency in one or more of the normal constituents. 

Anhydremia. — A deficiency in the normal fluid of the 
blood. 

Basophilic Granulation. — A peculiar granular degeneration 
of the red blood-cells which is noted in chronic lead-poisoning. 

Hydremia. — An excess of fluid in the blood. 

Leukocytosis. — An increase above the normal number of 
white bloocl-cells. 

Leukopenia. — A diminution in the number of white blood- 
cells. 

Lipemia. — The presence of an abnormal amount of free 
fat in the blood. 

Macroblasts. — Xucleated red blood-cells of more than nor- 
mal diameter. 

Macrocytosis. — The occurrence of numbers of abnormally 
large red blood-cells. 

Melanemia. — The presence of free pigment in the blood. 

Microblasts. — Xucleated red blood-cells of abnormally 
small size. 

Microcytosis. — The occurrence of numbers of abnormally 
small, red blood-cells. 

Normoblasts. — Xucleated red blood-cells of normal diam- 
eter. 



48 THE BLOOD. 

Oligochromemia. — A diminution in the normal amount of 
hemoglobin. This may occur either independently or coin- 
cidently with a diminution in the number of red blood-cells. 

Oligocythemia. — A diminution in the number of red bloocl- 
cells. 

Plethora. — An increase in the total quantity of blood above 
normal. 

Poikilocytosis. — This term is applied to the very irregular 
shape of the erythrocytes observed in certain pathologic condi- 
tions. 

Polychromatophilic Degeneration or Anemic Degeneration 
(Ehrlich). — This is an atypical staining reaction of the eryth- 
rocytes, the significance of which is not yet definitely deter- 
mined. 

Polycythemia. — An increase in the number of red blood- 
cells as compared with the fluid-content. 

VARIETIES OF LEUKOCYTES. 

1. Normal Leukocytes. — (a) Lymphocytes: These are 
derived from the lymph-glands, and appear as small, round cells 
about the size of a red blood-corpuscle, with a large, centrally 
located nucleus, and a small margin of protoplasm. The nu- 
cleus stains rather intensely with the nuclear stains (hematox- 
ylin, methylene-blue, and Ehrlich's triple stain). The proto- 
plasm is free from granules. 

(b) Large Mononuclear Leukocytes. — These cells are 
two or three times as large as a red blood-cell, with a large, 
usually oval nucleus which is generally eccentrically placed. It 
stains poorly with nuclear stains. There is a relatively large 
amount of protoplasm, which is free from granules. They are 
derived from the bone-marrow, and may be regarded as the 
parent type of the polymorphonuclear. 

(c) POLYNUCLEAR OF POLYMORPHONUCLEAR (NEUTRO- 
PHILIC Leukocytes. — These are recognized by their multiple, 
irregular-shaped, or bent nucleus. The nuclei stain very in- 
tensely, and the protoplasm is densely packed with neutrophilic 
granules. 

(d) Eosinophilic Cells.— These resemble the polymor- 



THE DIFFERENTIAL COUNT. 49 

phonuclear cells, except thai the small neutrophilic granules are 

replaced by coarse acidophilic (eosinophilic) granules. These 
granules are highly refractive, so that they can be readily recog- 
nized without staining. 

(e) Mast Cells. — These are cells of the polymorpho- 
nuclear type with marked basophilic granules, which are quite 
large, uneven, and irregularly distributed. They are not dis- 
tinctly stained by the triple stain. 

2. Pathologic Leukocytes. — (a) Mononuclear Neutro- 
philic Cells (Myelocytes) : These are large cells with a 
large, faintly-staining nucleus, differing from the large mono- 
nuclear cells of normal blood by the presence of neutrophilic 
granules in the protoplasm. 

(b) MonoxuclexVr Eosinophilic Cells (Ehrlich's 
Eosinophilic Myelocytes) : These cells are what their name 
implies, mononuclear eosinophiles. Very small cells of this type 
have been termed eosinophilic microcytes.' 

THE DIFFERENTIAL COUNT. 

The distinguishing characteristics of the various white 
cells, as brought out by any one of the differential stains above 
outlined, allow r the investigator to separate these into several 
groups, and thus to estimate their relative numbers which are 
usually expressed in percentage. The cells for a differential 
count can be enumerated without the aid of a mechanical stage, 
but this instrument of precision is a distinct aid in the per- 
formance of this procedure and should always be used when 
possible. Select a uniform and not too dense part of a stained 
smear, and adjust upon the mechanical stage so that the field 
presents cells in uniform arrangement without any overlapping. 
Xow, by means of the thumb-screws of the mechanical stage, 
the field is carried back and forth before the eye of the observer, 
so that the same part of the field is not brought into view more 
than once (this matter is very simple when a mechanical stage 
is used). As the parade of cells passes before the eye, the white 
cells are observed, classified, and the number jotted down. This 
is continued until not less than two hundred, and preferably five 
hundred or a thousand, cells have been counted. With the total 



50 THE BLOOD. 

number of cells counted known, and also the number of cells 
in each class recorded, it is a simple matter to calculate the 
percentage of each variety of cell in the specimen examined. 

The Normal Differential Count. — The normal leukocytic 
count may vary between 5000 and 10,000 white cells per cubic 
millimeter. The average normal leukocyte count is usually 
placed at 7500 leukocytes per cubic millimeter. 

The minimum and maximum number of cells for each 
type, estimated from the minimum normal (5000) leukocyte 
count, is given in the following table : — 

VARIETY. MINIMUM. MAXIMUM. 

Polymorphonuclears 3000 3500 

Small lymphocytes 1100 1500 

Large lymphocytes 250 350 

Eosinophiles 50 100 

Mast cells 5 25 

The minimum and maximum number of cells of each cell- 
type estimated from the maximum normal leukocyte count (10,- 
000). 

VARIETY. MINIMUM. MAXIMUM. 

Polymorphonuclears 6000 8000 

Small leukocytes 2200 3000 

Large leukocytes 500 900 

Eosinophiles 100 200 

Mast cells 10 50 

For the average normal standard for each cell-type we may 
adopt the following standard : — 

VARIETY. NUMBER. PER-CEXT. 

Polymorphonuclears 4875 65 

Small lymphocytes 1950 26 

Large lymphocytes 525 7 

Eosinophiles 75 1 

Mast cells 8 0.1 

LEUKOCYTOSIS. 

For convenience in the study of the various forms of leu- 
kocytosis, they may be divided into two classes : (a) Physiologic 
leukocytosis, (b) Pathologic leukocytosis. 

(a) Physiologic Leukocytosis. — The average number of 



LEUKOCYTOSIS. 51 

leukocytes, per millimeter of blood, is normally 7500. For chil- 
dren the average is slightly more. For weak and poorly nour- 
ished persons slightly less. The numbers of leukocytes in the 
peripheral blood of any individual vary from time to time. 
They (1) may be increased after a hearty meal, especially if it 
contains much proteid material. Physiologic leukocytosis may 
also occur (2) during pregnancy, particularly during the latter 
months of the condition; (3) in the new-born, and (4) after 
cold baths. 

In these so-called physiologic leukocytoses the increase does 
not usually exceed 30 per cent, of the normal, though in children 
it may be doubled. 

A liypo-leukocytosis is said not infrequently to precede a 
hyper-leukocytosis. 

(b) Pathologic Leukocytosis. — Many infections cause an 
increase in the number of white corpuscles in the peripheral 
circulation. Although the varieties of cells are the same as in 
health, the relative proportions are usually altered. In the most 
common form of pathologic leukocytosis, the percentage of lym- 
phocytes is diminished, while the polymorphonuclears are fre- 
quently increased from normal (65 per cent.) to 90 or 95 per 
cent. 

The polymorphonuclear leukocytosis occurs especially during 
inflammatory processes, and above all in those accompanied by 
purulent exudation. 

In certain infectious diseases, notably typhoid fever and 
uncomplicated tuberculosis, there is usually no increase in the 
number of white blood-cells. 

The origin of the extra leukocytes has not yet been de- 
finitely determined, as we do not know whether they are derived 
from the bone-marrow, lymph-glands, or from other tissues. 

Another form of leukocytosis is characterized by a relative 
increase in the number of eosinophilic leukocytes. This condi- 
tion of the blood is notably observed in bronchial asthma, trichi- 
nosis, and infections with other animal parasites. It is of in- 
terest to note that in these conditions there usually exist local 
collections of eosinophils at the seat of disease. Thus, in the 
walls of the bronchi and in the exudate in bronchial asthma, and 
about the embryos in trichinosis. 



52 THE BLOOD. 

The number of white cells in a pathologic leukocytosis not 
infrequently' reaches 20,000 to 30,000 cells per cubic millimeter, 
and has been known to reach the enormous number of 168,000 
(Grawitz). 

Leukopenia. — A diminution in the number of leukocytes in 
the peripheral blood occurs in a variety of conditions. It has 
been observed in cachexias, in intoxications, many anemias, and 
in some infectious diseases, notably in typhoid fever and in 
malaria. 

In leukopenia, as in leukocytosis, the relative proportions 
of the various varieties of white cells are usually changed. For 
example, in typhoid fever there is often a relative increase in 
the number of lymphocytes. 



THE ANEMIAS. 

Anemias are conveniently classified as primary when due 
to some unknown cause, and in which the blood-changes are, 
as a rule, of both a quantitative and a qualitative type. 

And as secondary, when the cause is known and when the 
blood-changes are usually of a quantitative type only. 

The Secondary Anemias. — Causes : Secondary anemia is 
observed after hemorrhage, during pregnancy, during chronic 
and constitutional diseases, and in poisoning, including that large 
and vague group of conditions comprising auto-intoxication. In 
chronic digestive disorders, malignant tumors, tuberculosis, syph- 
ilis, malaria, and in the different forms of helminthiasis. 

The Blood-Chaxges. — The chief blood-changes in sec- 
ondary anemia consist in a reduction of hemoglobin and a 
diminution in the number of red blood-cells. Mild forms show 
no other changes in the blood-picture. Severe forms show poi- 
kilocytosis, macrocytosis, and microcytosis. The extent to which 
these conditions are observed corresponds roughly to the severity 
of the anemia. 

Further the red corpuscles often manifest a change in reac- 
tion to the ordinary staining reagents. They stain poorly, un- 
evenly, and some parts of some cells refuse entirely to take the 
stain. This condition is termed polychromatophilia. This 



THE ANEMIAS. 53 

change is no indication of the grade of the anemia, as it is 

observed in the mildest forms of secondary anemia. 

The white cells in the simple anemias present nothing that 
is characteristic. 

Summary. — The essential blood-changes in secondary, or 

simple, anemias consist in a diminution in the hemoglobin per- 
centage, and in the number of red blood-cells. The red cells 
may show polychromatophilic degeneration and poikilocytosis. 

The Primary Anemias. — General Considerations : Un- 
like the secondary anemias, the blood-changes in primary an- 
emias, besides showing any or all of the modifications observed 
in the former, present very striking and characteristic alterations 
in the white blood-cells. 

Progressive Pernicious Anemia. — In contrast to the process 
in simple anemia, in progressive pernicious anemia, blood-degen- 
eration in certain portions of the blood-making organs, notably 
in the bone-marrow, takes place in a manner different from the 
physiologic. Consequently, in the blood-formative organs and 
also in the circulation, we note cells often in great numbers that 
are never seen in the normal blood. These pathologic elements 
are present in embryonal life ; so in pernicious anemia we speak 
of the reversion of blood-formation to the embryonal type. 

The Blood-Changes. — In a typical, well-defined case of 
pernicious anemia, the first glance at a well-prepared stained 
specimen of blood is sufficient to separate it immediately from 
the class of simple anemias. We find that a large number of 
erythrocytes have a diameter greater than normal (megalocytes : 
diameter 15 to 16 microns). These cells by their staining show 
a great richness in hemoglobin. Careful search always shows 
some megaloblasts, i.e., nucleated (embryonal) red corpuscles. 
Xormoblasts and microcytes are also in evidence. Other changes 
usually present are poikilocytosis, polychromatophilia, and gran- 
ular degeneration. 

The red corpuscles are always notably decreased, and may 
be less than 20 per cent, of the normal. The percentage of 
hemoglobin is also diminished, but practically always in a less 
degree than the red cells. On account of this condition the 
color-index is very frequently above 1.0. 



54 THE BLOOD. 

CHLOROSIS. 

Blood-Changes. — The blood when drawn flows freely from 
the puncture, and is markedly watery. Hemoglobin estimation 
shows a decided reduction in the percentage of hemoglobin with- 
out a corresponding deficiency in the number of red blood- 
cells. The color-index is therefore low, below 1.0, in contrast 
to the high color-index observed in pernicious anemia. This 
particular characteristic is not unique in chlorosis, as other forms 
of anemia may also show it. 

Morphologically we find striking changes in the erythro- 
cytes (many appear as macrocytes), which are pale and are 
without a distinctly pronounced central umbilication — the cells 
appear swollen. The particular cells have been designated "chlo- 
rotic" blood-corpuscles. Severe cases shows poikilocytosis, and 
nucleated red blood-cells. 

Polychromatophilia and granular degeneration, which are 
the genuine phenomena of degeneration, are not observed. 

The condition of the leukocytes is not uniform. The cells 
themselves do not show any characteristic changes in this dis- 
ease. 

The blood-plaques appear in markedly increased numbers, 
so that many groups of these cells appear in every field of the 
fresh preparation. 6 These are often very evident in the stained 
preparation, especially if a basic stain, such as methylene-blue, 
has been used. 

LEUKEMIA. 

Changes in the Number of Erythrocytes. — The blood from 
a marked case of leukemia is distinctly watery ; in extreme cases 
it may be a whitish-red as it emerges from the puncture; this 
is owing to the great increase in white elements. 

The microscopic examination of the fresh blood in estab- 
lished cases, shows even without counting the enormous increase 
in the white blood-cells. The count of the white corpuscles 
shows 100,000 to 500,000, or even more, to the cubic millimeter. 

In some cases of leukemia no noteworthy change in the 
erythrocytes, either in number or appearance, occurs. As a rule 



6 E. Grawitz: Modern Clin. Med., p. 327., 



LEUKEMIA. 55 

they are decreased to about half the normal number. Beside 
the diminution in these cells the blood contains varying numbers 
of normoblasts, and more rarely megaloblasts. 

The amount of hemoglobin is diminished, but the coloring 
of the individual corpuscles need not be diminished (no altera- 
tion in the color-index). 

Changes in the Blood-Plaques. — An increase in the number 
of the blood-plaques has been noted in a number of cases. 

Blood Morphology. — The diagnosis of leukemia can fre- 
quently be made without further consideration in cases which 
show an extraordinary increase in the number of white elements, 
but in doubtful cases the presence of the condition can only be 
determined by an exact examination of the morphology of the 
leukocytes. 

Besides noting the great increase in the number of white 
cells, the polymorphonuclear elements will usually be found the 
most numerous. 7 In other cases the increase is chiefly among 
the lymphocytes. We may therefore differentiate two forms of 
leukemia. 

1. Lymphatic Leukemia. 2. Leukocytic Leukemia. 

Since the source of the lymphocytes is from the spleen and 
from the lymph-glands, and that of the leukocytes from the 
bone-marrow, the common designation of lymphatic leukemia 
(lymphemia) and myelogenous leukemia (myelemia) may be ap- 
plied to these two basic forms of leukemia. 

As, however, the bone-marrow normally produces typical 
lymphocytes, and as it has been demonstrated in rare conditions 
that an overflooding of the blood with lymphocytes may occur 
through proliferative changes in the bone-marrow without en- 
largement of the spleen or lymph-glands (myelogenous lymph- 
emia). It seems preferable to apply the less prejudicial division 
of the leukocyte forms into "lymphatic leukemia" and "leuko- 
cytic leukemia" (W. von Leube). 

Differentiation. — 1. Lymphatic Leukemia: The blood- 
picture is conspicuous for its great preponderance of large and 
small lymphocytes, in comparison to the leukocytes. Nucleated 
red cells (normoblasts) and megaloblasts, although present in 



7 W. von Leube: Modem Clin. Med., p, 349. 



56 THE BLOOD. 

this form of leukemia, are by no means as abundant as in the 
leukocytic form. 

2. Leukocytic Leukemia. — This is by far the most com- 
mon form of leukemia. It may easily be differentiated from 
the preceding by the entirely different blood-picture. The in- 
crease in the white elements is usually very marked. Here, how- 
ever, the polymorphonuclear leukocytes are greatly increased 
in the microscopic blood-picture. Xeutrophiles and eosinophiles 
are absolutely always increased (Ehrlich). There is also an 
increase in the mast cells, which may be twice as numerous as 
the eosinophiles. Their determination is of the greatest im- 
portance, since a marked increase in mast cells is observed only 
in this disease (von Leube). 

The phenomena which especially characterize leukocytic 
leukemia, and which result from changes in the bone-marrow 
(myelogenous leukemia) are the occurrence of neutrophilic 
and eosinophilic myelocytes. These may be present in enormous 
numbers (up to 100,000 per cubic millimeter). The first sight 
of such a picture simulates the blood-picture of acute lymphemia 
with its large mononuclear cells. 

The myelocyte is a large mononuclear cell with an irregular 
nucleus, surrounded by a considerable amount of protoplasm, in 
which are either neutrophilic or eosinophilic granulations. # Be- 
sides these immature leukocytes (myelocytes), immature forms 
of erythrocytes, also originating in the bone-marrow, are found 
in the circulating blood-stream of patients suffering from leuko- 
cytic leukemia. There are also normoblasts and occasionally 
megaloblasts. 

SPECTROSCOPIC EXAMINATION. 

Solutions of hemoglobin and its derivative compounds, 
when examined with the spectroscope, give distinctive absorption 
bands. 

The Spectroscope. — Light, when made to pass through a 
glass prism, is broken up into its component rays, giving the 
play of rainbow colors known as the spectrum. A spectroscope 
is an apparatus for producing and observing the spectrum. In 
brief, the apparatus consists of a base or stand, two horizontal 



SPECTROSCOPIC EXAMINATION. 57 

tubes, and a prism arranged to take the lighl coming from one 
and to pass it into the other. 

Light falls upon the prism through one tube known as the 
"collinator tube." A slit at the end of this tube admits a nar- 
row ray of light which, by means of a convex lens in the other 
end of the tube, is made to fall upon the prism with its rays 
parallel. In passing through the prism the ray of light is dis- 
persed by unequal refraction, giving the spectrum. The spec- 
trum thus produced is examined by the observer through the 
other tube, which is a telescope. ^Yhen the telescope is properly 
adjusted the rays entering from the prism produce a clear pic- 
ture of the spectrum. If the light used is lamplight, then the 
spectrum will be continuous, the colors gradually merging one 
into the other from red to violet. If sunlight is used the spec- 
trum will be crossed by a number of narrow, dark lines known 
as "Fraunhofer lines." The position of these lines in the solar 
spectrum is fixed, and the more distinct ones are designated by 
the letters of the alphabet, A, B, C, D, E, etc. 

If, while using artificial light or the solar spectrum, a solu- 
tion of any substance which gives absorption bands, is placed 
in front of the slit so that the light is obliged to traverse it, then 
the spectrum, as observed in the telescope, will show one or more 
narrow or wide, black bands, which are characteristic of the 
substance examined and w T hich constitute its absorption spec- 
trum. The position of these bands may be located by describing 
their relation to the Fraunhofer lines. 

While the cost of the spectroscope may prohibit its use by 
the general practitioner, it is to be found in many laboratories, 
and its use in certain cases, particularly in poisoning, is abso- 
lutely essential, since it enables the student to arrive at definite 
conclusions which cannot be reached in any other way. 

For ordinary investigation the pocket spectroscope is all 
that is required. The detection of CO-Hb and of methemo- 
globin, the first occurring in carbonic-acid poisoning, the second 
in the various forms of intoxication, particularly with chlorate 
of potash. The smallest amount of hemoglobin or its deriva- 
tives may be demonstrated with certainty by this means in cases 
where ocular examination would leave uncertainty. 

The determination of CO-Hb (carbon-monoxid hemoglobin) 



58 THE BLOOD. 

may present difficulty on account of the similarity of the spec- 
trum to that of O-Hb (oxyhemoglobin). The differentiation 
is only certain when we observe that the line of CO-Hb does not 
appear on the addition of reducing agents — for example, ammo- 
nium sulphate. The CO-Hb is a strong combination, while, on 
the other hand, the O-Hb is changed by the reagent to that of 
reduced hemoglobin. 



BACTERIOLOGIC EXAMINATION. 

Both diagnosis and prognosis may, in some cases, be helped , 
by a bacteriologic examination of the blood. As there are quite 
a number of diseases in which the pathologic agents enter the 
blood, where diagnoses may thus be corroborated by a simple 
microscopic investigation of the blood, as in relapsing fever, 
malaria, and syphilis. In other cases, because of the small total 
number of organisms, or of the presence of others due to con- 
tamination, it is advisable to proceed by the more tedious but 
more certain bacteriologic methods. 



THE CUNICAL VALUE OF BLOOD CULTURES.* 

"Blood-cultures are most likely to be of value in the fol- 
lowing diseases : Typhoid fever, pneumonia, other forms of pneu- 
mococcic infections, and the whole group of septic cases in which 
the sepsis is associated with wounds, pelvic diseases, abortion, 
puerperal infection, endocarditis, and local diseases of the throat 
and many other regions. 

"In infections with typhoid bacillus, the pneumococcus, 
staphylococcus, and streptococcus, the results are usually posi- 
tive. 

"In a case of typhoid the examinations along bacteriologic 
lines are most likely to be successful between the latter part of 
the first and the middle of the third week. In sepsis due to the 
pyogenic cocci positive results are obtained in the majority of 
cases. 

"Blood-cultures may clear up doubt in the presence of 

8 Edsall: quoting Longrscope and Evans. 



THE SPIROCHETE PALLIDA. 59 

typhoid fever or pulmonary tuberculosis. They may also be of 
considerable use in obscure infections in childhood/' 

The Specimen. — In the preparation of blood-cultures it is 
advisable to puncture a vein in the arm under strict aseptic 
precautions to withdraw one or two cubic centimeters of blood, 
and to proceed immediately to prepare cultures or proceed with 
animal experiments. 

Relapsing Fever. — The diagnosis of this disease is only 
positive when the spirilli are found in the fresh blood. As a 
rule this examination is easy, for the peculiar curved form of 
the spirillum precludes confusion with other organisms. Their 
numbers are usually large and their detection facilitated by their 
motility. In addition they stain readily with the various aniline 
colors (as carbol-fuchsin). 

Malaria. — In this disease the microscopic examination is 
of extreme practical importance. The organisms may be readily 
detected in fresh blood at the bedside w T ith the aid of a good 
microscope. The dried stained specimen, which is usually the 
more practical to procure in practice, will serve to detect them 
even when in but small numbers. 

The specimen for examination is best taken in the afebrile 
period, shortly before the expected paroxysm. Previous to this 
the administration of quinine should be withheld for as long a 
period as possible. 

THE SPIROCHETE PALLIDA. 

Description of the Spirochete. — The spirochsete is four- 
fourteenth microns in length, and at most only one-quarter of a 
micron in breadth; its form is that of a corkscrew with six to 
fourteen or even twenty-four turns. It is in active motion when 
examined in the fresh, and may show this activity for days in 
salt solution, when the tissues are kept at 20° to 27° C. (Beer). 

Method of Obtaining a Smear (Roescher). — The surface of 
the sore is cleaned with alcohol, irrigated with salt solution, and 
dried. It is then scraped with a needle, care being taken to 
avoid drawing blood, but bringing out a good deal of serum. 
Smears are made from the serum, and in it the spirochete will 
be found. Serum may be drawn from an enlarged gland by 



go THE BLOOD. 

means of a hypodermic needle and the smears made from this if 
desired. 

The Giemsa Method (for preparation of stain see Appen- 
dix). — The films are fixed in methyl, or ethyl alcohol, and dried 
with blotting paper. The stain is then diluted with distilled 
water in the proportion of one drop to 1 cubic centimeter of 
water, and then poured on the specimen. For satisfactory results 
the stain should remain on the specimen for from one to twenty- 
four hours. The time for staining may be materially shortened 
by adding a few drops of a 1 iooo solution of potassium carbonate 
to the diluting water. By this expedient the spirochete may be 
demonstrated in fifteen minutes, though for the best results an 
hour's contact with the stain is advised. 

Method of Oppenheim and Sachs. — The smears are dried in 
the air and then, without previous fixation, are flooded with an 
alcoholic solution of carbol-gentian- violet (for preparation of 
stain see Appendix). After flooding the preparation it is 
warmed until it steams for a few minutes only; it is then 
washed gently in running water, dried, and examined. The 
spirochete are stained blue. 

Method of F. C. Wood (fifteen minutes).— The smears are 
fixed in strong methyl alcohol and dried with blotting paper. 
The following stains are then applied in succession : — 

First, a few drops of a %ooo watery solution of yellow 
water soluble eosin are spread over the smear by means of a 
pipette. Second, four or five drops of a %ooo aqueous solution 
of methylene azur II. The two colors are thoroughly mixed by 
spreading over the smear and by rocking the cover-glass. This 
mixture is allowed to act for ten minutes. The stain is then 
washed off by a strong stream of distilled water, the preparation 
dried between blotters, mounted, and examined by an oil-immer- 
sion. For mounting use xylol-dammar, as the ordinary Canada 
balsam bleaches these preparations. Usually any precipitate 
which forms can be removed with the stream of water. If espe- 
cially clean and clear specimens are desired, the precipitate may 
be removed by momentary immersion in 95 per cent, alcohol, 
which should be immediately washed off with distilled water. 
By this method the spirochete appear a light carmine color. 



AGGLUTINATION REACTIONS. Gl 

AGGLUTINATION REACTIONS. 

The Phenomenon. — We owe a valuable diagnostic method 
to the knowledge derived from the law of immunity. We know 
that in the natural infections of man, as well as by animal ex- 
periments, peculiar changes occur in the blood-serum under the 
influence of specific bacteria or their soluble toxins. Among 
these altered sera we differentiate two classes: 1. The antitoxic 
sera. 2. The bactericidal sera. The first variety is represented 
by the serum containing the diphtheritic antitoxin; the second 
by the energetic bacteriolytic action of the blood of convalescents 
from cholera. 

As another variety of the latter, the group of agglutinating 
sera may be included ; these occur during the course of and after 
recovery from many infections. We are most familiar with this 
condition after typhoid fever. 

The Widal Reaction. — Definition: Agglutination in the 
bacteriologic sense refers to the clumping and sedimentation of 
micro-organisms from homogeneous suspension by the action of 
a serum. This is the so-called Gruber-Widal reaction. 

Xormal serum may agglutinate many bacteria, as typhoid, 
colon, pyocyaneus, and dysentery, but not the streptococcus 
and a number of other organisms. This agglutination only 
occurs in low dilutions, though the typhoid bacillus has been 
known to agglutinate in as high a dilution as %o> a point to be 
remembered in practical diagnosis. 

Xearly all the bacteria, even the non-pathogenic, will pro- 
duce agglutinating serum when injected, but not all have this 
power equally. 

Technic. — Two methods : 1. The macroscopic, or naked 
eye, observation of the clumping and sedimentation of a homo- 
geneous suspension of bacteria in test tubes. 2. The microscopic 
observation of the clumping of the organisms when the bacteria 
are mixed with serum and mounted in a hanging-drop prepara- 
tion. 

For the hanging-drop preparation it is necessary to have a 
slide with a saucer-shaped depression in the middle of one sur- 
face. A drop of the solution to be examined is placed in the 
center of a cover-glass, which is then placed drop-side down over 



g2 THE BLOOD. 

the depression, and the edges sealed by vaseline or paraffine. In 
this there is ample room for the motile organisms to swim about, 
and the loss of motility incident to the agglutination is readily 
observed. 

The Culture. — When the organism to be tested grows rap- 
idly, it is absolutely necessary to use a fresh, young culture, one 
which has been grown at room, not incubator, temperature, on 
agar or in bouillon for from eighteen to twenty-four hours. If 
the agar culture is used the bacteria may be washed from the 
surface by pouring into the tube 5 to 10 cubic centimeters of 
normal salt solution, and shaking vigorously. The resulting sus- 
pension is then ready for use. A homogeneous suspension is 
absolutely necessary to successful work. Accidental agglutina- 
tion from insufficient dilution, must always be suspected and 
guarded against. 

The Serum. — A blister may be used. A cantharides plaster 
from one-half to three-quarter inch square is placed on the ab- 
dominal skin and covered with a dressing. In about fourteen 
hours remove the serum with a sterile hypodermic syringe, or 
0.5 to 1 cubic centimeter of blood can be drawn in the usual 
way from the lobe of the ear or the tip of the finger, and the 
serum removed after coagulation has occurred, or the serum 
may be immediately separated in a suitable tube in the centri- 
fuge. 

The custom of drying a few drops of blood on a piece of 
paper, as practiced in many municipalities where the specimens 
are sent by mail to a central laboratory, is very convenient, and 
in general practical, but it precludes the possibility of using 
definite dilutions, because the bacteriologist does not know the 
precise amount of serum or blood used. (See Illustration of 
Philadelphia Test Blank.) 

Serum Dilutions. — Ordinary serum of dilution, 1 to 10 
or less, will often agglutinate the typhoid bacillus and other 
organisms, while higher dilutions, 1 to 30 or 1 to 60, fail to 
act. The serum of the typhoid patient or convalescent, however, 
rarely fails to agglutinate in these dilutions. 

It is usually held that dilution of 1 to 40 or 1 to 50 is 
sufficiently high to eliminate the possibility of false agglutinat- 
ing reactions by normal serum, and at the same time sufficiently 



WIDAL REACTION. q$ 

low to permit of agglutination of nearly all the sera of typhoid 

cases. 

Loop Dilution. — A convenient method of measuring small 
amounts of culture and serum is to measure by means of a fine 
wire loop, such as used in bacteriologic work. One loop full of 
serum placed on a slide and diluted with ten loops full of water, 
will give a dilution of 1 to 10. By this means minute quanti- 
ties of serum can readily and quite accurately be reduced to the 
desired dilution. 

If agglutination occurs in a microscopic preparation, we 
note, with the aid of the high power, that, in the course of 
from fifteen minutes to an hour as the micro-organisms swim 
about, a few which come in contact have a tendency to remain 
in this position. In the course of a few more minutes other 
organisms join this group and other groups form throughout 
the field ; motility becomes gradually less until it ceases entirely 
in the typical reaction. The complete change occurs in from 
six to eight hours. Xo less than five bacteria must become 
permanently agglutinated to constitute a positive reaction. The 
test is most decisive when large masses of permanently agglu- 
tinated organisms are formed which can be seen under the lower 
powers. 



IV. 

OPSONIC METHOD. 



The Theory. — Two elements come into consideration in 
the protection of the organism against invading micro-organ- 
isms. The leukocytes with their digestive ferments constitute 
one of these elements; the antibacterial substances in the blood 
fluids constitute the other. 

(a) Leukocytes. — The leukocytes come into consideration 
in connection with the resistance of the bacterial infection, by 
virtue of the fact that they are capable of ingesting bacteria and 
of disintegrating these by intracellular digestion. 

We may usually distinguish between "spontaneous" and 
"induced" phagocytosis. 

"Spontaneous" phagocytosis is the process of ingestion and 
destruction which occurs in different media, as physiologic salt 
solution. It is a slow process and the bacteria ingested are in 
moderate numbers. It is also irregular, in that the number 
ingested of different species of bacteria varies. 

Strikingly different from such spontaneous phagocytosis is 
the "induced" phagocytosis, which comes under observation when 
leukocytes are brought into contact with bacteria which have 
been, or actually are at the moment, subjected to serum action. 
The "induced" phagocytosis which occurs under these conditions 
is distinguished by the following facts : — 

1. It is an exceedingly rapid process. 2. With hardly an 
exception every adult leukocyte is phagocytic. 3. When the 
supply of organisms is unrestricted the leukocytes will ordina- 
rily fill themselves to absolute repletion. -A. The leukocytes will 
continue to digest bacteria in a concentration of salt solution 
which would entirely suppress "spontaneous phagocytosis." 1 

(b) Antibacterial Elements in the Blood. — The antibac- 



1 Sir A. E. Wright: Jour. Araer. Med. Asso., Aug. 10-17, 1907. 

(64) 



GENERAL CONSIDERATIONS. 65 

terial elements which arc here in question are of ''bacterio- 
trophic" properties, in that they turn toward and enter into 
combination with the elements of the bacterial body. Our knowl- 
edge of the modifications which are effected in the bacterial 

body under the influence of tin 1 bacteriotrophic substances in the 
blood-fluids, is far from complete. 

This bacteriotrophic action may manifest itself in various 
ways : 1. The bacteria may be killed without being dissolved. 2. 
The bacteria may not only be killed, but also dissolved. We may 
group these activities as bacteriocidal and bacteriolytic effects. 
3. The bacteria may be so altered as to agglutinate in the pres- 
ence of salt (agglutination reaction). 4. The bacteria may be 
so affected as to be readily ingested by the phagocytes (opsonic 
effect). 

These effects can be experimentally proven. We may as- 
sume then, that we have in the blood-fluids, in addition to bac- 
teriocidal substances (bacteriocidins), also bacteriolysins, agglu- 
tinins, and opsonins. 

Of these four varieties of bacteriotrophic substances, the 
opsonins appear to be the most important. We are justified in 
ascribing to them predominating importance, because it can be 
shown that the opsonic effect is exerted either by the normal 
or immune blood of every species of bacteria, while the bacterio- 
cidal and bacteriolytic activities are exerted among pathologic 
bacteria, apparently only on the typhoid bacillus and the cholera 
vibrio. The opsonic effect is of further importance, because the 
effect can be accurately measured (Wright). 

General Considerations. — The opsonic substances are essen- 
tial to induced phagocytosis. They do not act upon the leuko- 
cyte (as a stimulant to the cell), but combine with the micro- 
organisms, thus preparing them for the leukocytes. Hence the 
term Opsonin, from opsono, I cater for. The opsonins have 
been shown to be almost destroyed by heating to 60° C. for a 
short time. The opsonic power of the blood gradually disap- 
pears on standing, losing about 50 per cent, of its activity in 
five or six days. 

When a condition of immunity is conferred upon a patient 
infected with staphylococci, the opsonic activity of the patient's 
blood is greatly augmented. 



QQ OPSONIC METHOD. 

It is possible that there are present in the blood-plasma 
many varieties of opsonins, each concerned with combating one 
particular kind of microbic invasion. 

The leukocytes of healthy or of diseased persons seem 
equally active when brought in contact with the same serum; 
hence the amount of opsonins present in the blood of an indi- 
vidual determines, according to the opsonic theory, his sus- 
ceptibility to bacterial invasion. 

The Techxic. — To measure the resistance of a patient 
to a bacterial invasion, in other words, to find his opsonic index 
toward a certain micro-organism, special technic has been devel- 
oped, the salient points of which will be briefly described. 

Bacterial Emulsions. — Young cultures of rapid-growing 
bacteria, such as staphylococci, pneumococci, gonococci, and colon 
bacilli, are inclined upon agar-agar for a period of not longer 
than twenty-four hours in the incubator. They are then washed 
off with normal salt solution. After this has sedimented for 
a short time the upper whitish layer is removed with a pipette 
into a small centrifuge tube, and the larger clumps of bacteria 
removed by rotating for a few minutes. The supernatant layer 
which is still whitish and opalescent, is called the bacterial emul- 
sion. An emulsion, if suitable for work, should contain the 
germs well separated and evenly distributed. 

In preparing emulsions of tubercle bacilli, the culture is 
heated, then ground in a mortar with salt solution until finely 
divided, after which the mixture is centrifugated and the 
cloudy layer of emulsion removed. 

Determination of the Strength of the Emulsions. — The 
method of Wright consisted in counting the actual number of 
germs in a measured amount of emulsion; the process is both 
difficult and tedious. McFarland and L'Engle have devised an 
apparatus for estimating the strength of bacterial emulsions, 
which is termed a "nephelometer." This consists essentially of 
a series of sealed glass tubes containing various strengths of 
barium sulphate solution. These serve as standards for com- 
parison of the density of prepared bacterial emulsions. For the 
best results it has been found that the tube corresponding to 
the 5-per-cent. solution of BaS0 4 is the best. (For complete 
description of the nephelometer see end of this section.) 



METHOD OF OBTAINING THE OPSONIC INDEX. 67 

The permanency of the emulsions varies a great deal. Sus- 
pensions of gonocoeci should be used at once. Those of staphy- 
lococci within two days, while the emulsion of the tubercle bacilli 
may be kept indefinitely. 

Washed White Corpuscles. — The finger or lobe of the ear 
is prepared and punctured as for the blood-count, and the blood 
as it exudes is allowed to fall directly into normal salt solu- 
tion (0.85 per cent.) containing about 1 per cent, of sodium 
citrate. This decalcifies the blood and prevents it from clotting. 
The corpuscles are then completely precipitated by the centri- 
fuge, and then alternately washed with salt solution and cen- 
trifugated until all trace of the sodium citrate is removed. After 
the final centrifugation the salt solution is decanted and the thin, 
upper layer of sediment, the "leukocyte cream," consisting for 
the most part of washed white corpuscles, is removed. 

The Serum. — The blood is obtained in the usual way and 
collected in small glass tubes suitable to place in the centrifuge. 
These are rotated and the serum separated. In obtaining the 
normal serum care must be exercised to select a healthy subject, 
or, what is better, to obtain a mixed specimen from a number 
of subjects. This is "pool serum." 

METHOD OF OBTAINING THE OPSONIC INDEX, 

Special capillary pipettes, graduated in three equal divi- 
sions, are required for mixing the washed corpuscles, the pa- 
tient's serum, and the bacterial emulsion. These are taken up 
into the tube in equal amounts and immediately blown out upon 
a microscope slide and thoroughly mixed. This mixture is again 
taken up into the tube and sealed in with the aid of a small 
Bunsen flame. The sealed tube is then transferred to an incu- 
bator of the ordinary type, or into an opsonic incubator as pro- 
posed by Freeman, and allowed to remain at a temperature of 
37° C. for exactly fifteen minutes. 

A second tube is prepared in the same way, only here normal 
or "pool" serum is substituted for that of the patient. 

At the expiration of fifteen minutes smears from each tube 
are made upon cover-glasses and dried. These may be stained by 
any of the differential staining methods described in the section 



68 OPSONIC METHOD. 

on the blood. From the stained specimen a good field is selected 
(one containing a large number of leukocytes). The number 
of germs contained in the first fifty or one hundred cells is 
counted, and the total divided by the number of cells counted. 
This gives the the average number of bacteria per cell. Organ- 
isms which do not focus simultaneously with the cells should be 
excluded, as probably being extracellular. If the average num- 
ber of bacteria per cell in the mixture containing normal serum 
was two, then the opsonic index for the normal serum is two. 
If the number of bacteria averages one for an equal number of 
cells in the mixture containing the patients serum, then the 
opsonin index for that particular germ would be one-half. 

Wright and his pupils, as a result of numerous observations, 
have classified diseases due to bacterial invasion as follows : — 

1. Diseases in which the bacterial process is strictly local- 
ized or shut off from the lymph-channels, as furunculosis, lupus, 
and tuberculosis, and in fact almost any chronic infection. The 
opsonic index in these cases is constantly below normal, owing 
to the absence of immunizing stimuli emanating from the com- 
pletely walled-off infective process. 

2. Diseases in which the bacterial process is but loosely 
shut off, especially from the lymph-channels. In these, which 
are generally acute infections, immunizing agents from the bac- 
teria, from time to time, get into the circulation: thus the 
opsonic index may be normal, above or below normal, at vari- 
ous times. A good example of this class is tuberculosis of the 
joints. 

3. Diseases in which the bacterial infection is in the blood- 
stream. Here the opsonic index is usually above normal. To 
this class belong the infectious diseases : septicemia, endocar- 
ditis, etc. 

4. Normal individuals not subject to any bacterial infection 
present a constant opsonic power to the various pathogenic bac- 
teria. 

For opsonic treatment of bacterial diseases Wright and 
Douglass lay down the following rules: — 

1. Isolate in pure culture the causative micro-organisms. 

2. Estimate the opsonic power of the patient's blood to that 
micro-organism. 



THERAPEUTIC APPLICATION. 69 

3. If the opsonin index is at or below normal, as com- 
pared with norma] or "pool" serum, prepare and standardize a 
vaccine from this micro-organism. 

4. Inoculate the patient with this vaccine in appropriate 
doses at proper intervals, as shown by systematic examination of 
the opsonic content of the patient's blood. 

Reflection will make clear that the success of the immu- 
nizing process must depend, in the first place, on the power of 
the immunizing response which the human organism may pos- 
sess with respect to the particular bacterial infection or intoxi- 
cation process which is in question, and, in the second place, 
on (a) the composition of the vaccine and (b) the dosage and 
method of administration. 



STANDARDIZATION OF THE VACCINE. 

The emulsions, with the exception of the tubercle bacillus, 
consist of dead cultures of the organisms (killed by heating). 
The amount of heat for sterilization will vary for different or- 
ganisms: 60° C. or slightly less, for about thirty minutes, is 
usually sufficient. Cultures from the sterilized vaccines are then 
made to demonstrate that the culture is completely killed. The 
tubercle vaccine is the new tuberculin, "Koch." It is an opales- 
cent fluid containing the active principle of the tubercle bacillus 
obtained according to the method of Koch, and may be pro- 
cured in the open market. 

THERAPEUTIC APPLICATION. 

To successfully inoculate the prepared vaccine, it is neces- 
sary to have a pure culture of the organism, preferably grown 
from the patient's own strain of organisms. The patient's op- 
sonic index to this particular germ is then taken. The patient 
is then given an initial injection of the standardized bacterial 
emulsion, and his opsonic index watched. In a short time after 
the inoculation the opsonic index will be found lower than it 
was originally. This is the so-called "negative phase." Sooner 
or later, from a few hours to a few days, the opsonic index rises 
above the starting point. This is the "positive phase." The 



70 OPSONIC METHOD. 

amount of opsonins in the blood now remains stationary for a 
while, then gradually diminishes. As soon as this beginning 
diminution is noted, a second injection of vaccine is given, which 
is again followed by a negative phase, though less pronounced 
than before. Following this a positive phase occurs which even- 
tually reaches a higher level than the first. Thus the injections 
are repeated from time to time, according to the opsonic index of 
the patient, until the opsonic index in its positive phase reaches 
a normal or even higher level. In other words, if the injections 
are given properly (never in a negative phase), the patient's tis- 
sues are stimulated to an increased production of opsonins. The 
destruction of bacteria in the system is greatly increased, and 
the patient eventually recovers from the infection. 

In the study of this process it will be noted that increased 
phagocytic response is associated with successful immunization, 
and that this phagocytic response is dependent upon an increase 
in the opsonic power of the blood-fluids and not on an increased 
capacity for spontaneous phagocytosis on the part of the white 
corpuscles (Wright). 

When a dose of vaccine, which is only just sufficient to pro- 
duce a result, is administered, the negative phase is elided and 
there is registered only a positive phase. When an excessive dose 
of vaccine is administered (one which produces severe consti- 
tutional symptoms) the negative phase is proportionately accen- 
tuated and prolonged. 

The bacteriotrophic blood-power may be reduced for a 
period of weeks, and the advent of the positive phase in such a 
case awaited in vain. Immoderate negative phases due to over- 
dosage, rarely come under observation except in the case of 
patients who are new to inoculation. In case a prolonged nega- 
tive phase is produced by inadvertently administering an over- 
dose, the desired raise can practically always 'be obtained by re- 
inoculating as soon as all the constitutional symptoms have dis- 
appeared (Wright). 

It is a proper principle of dosage in any series of inocula- 
tions, never to advance to a larger dose until it has been ascer- 
tained that the dose which is being employed is too small to 
evoke an adequate immunizing response (Wright). 

A dose of vaccine may be adjudged too small as soon as it 



DIAGNOSIS OF OBSCURE LOCALIZED INFECTIONS. 71 

has been ascertained thai the inoculation is not followed by a 
negative phase, or when the negative phase is not very well 
marked and is of very short duration (Wright). 

A whole array of observations point to the local production 
of bacteriotrophie and vaccinotrophic substances generally at the 
Bite of inoculation. It has also been noted that local immunity 
may be acquired and retained apart from the acquirement or 
retention of general immunity. 



DIAGNOSIS OF OBSCURE LOCALIZED INFECTIONS. 

Localized infections can be made to produce auto-inocula- 
tion by massage or by forced respiratory efforts in the case of 
pulmonary tuberculosis. 

Auto-inoculation associated with measurement of the op- 
sonic index is of value in the preliminary diagnosis of bacterial 
infections. When inoculation by this means can no longer be 
induced (which originally was possible), we may conclude that 
the focus is extinct, and visa versa. 

Conditions which Present the Simplest Problems. — (a) 
Penetration of a single species of micro-organisms into the body. 
Examples: Tubercle bacilli in lymph-nodes, furunculosis. In 
the treatment of these cases successful results are accom- 
plished in a few days. 

Tuberculin is a valuable aid to diagnosis in early tubercu- 
losis, but when not properly used is dangerous, and in inex- 
perienced hands may bring disaster. 

The opsonic index may be of great aid in the early recog- 
nition of tuberculosis, especially where tuberculin cannot be 
given on account of temperature or prostration. The use of the 
opsonic index may give some indication for the control of the 
dose of tuberculin used for diagnosis as well as for its thera- 
peutic effect. 2 

Grace Calvert thus sums up our present knowledge concern- 
ing the value of the opsonic index in pulmonary tuberculosis : — 

1. In mild, early cases, if opsonic index is above normal, it 
is a favorable period of the disease. 



2 Rotch and Floyd: Jour. Amer. Med. Asso., Aug. 24, 1907. 



72 OPSONIC METHOD. 

2. In acute cases it fluctuates greatly from day to day. 

3. In chronic cases it is below normal. 

(b) Ulcerative types of infection as in the breaking-down 
of the deeper tissues. This is not more difficult to treat than 
the preceding, except where secondary infection has occurred. 

(c) Infections of the skin: (a) Those which are dry, scaly, 
and non-vascular ; these are extremely intractable, (b) Those 
where the skin is vascular or where there is purulent infection, 
as in acne and sycosis. This form is very tractable to vaccine 
therapy. 

H. E. Yarney 3 reports 9 cases of acne in which no other 
method of treatment was employed. Six cases were cured. One 
discontinued treatment; two required other treatment in con- 
junction with vaccination. 

A number of cases of furunculosis were cured with no 
recurrence in some months. 

In two cases of sycosis of two and six years' standing, the 
first was not affected after prolonged treatment; the other was 
speedily cured. 

(d) Infections of mucous membranes and glands and ducts 
connected therewith. These are readily influenced by bacterial 
vaccines when in the antrum, the nose, nasal sinuses, dental 
alveoli, salivary glands ; also in infections of the intestines and 
gall-bladder, different infections of the uterus, urinary bladder, 
and urethra. Mixed infections add to the difficulty of treating 
these cases. 

(e) Mixed infections. In a few instances the extinction 
of one species of bacterium by inoculation may lead to extinction 
of others which may be present. Such an event is extremely 
exceptional. More success follows efforts to immunize the pa- 
tient against the different infections. This is particularly the 
case in lupus, cystitis, and in endometritis. 

(f) Generalized infections. The vaccine method has favor- 
ably influenced some cases of Malta fever, and cures have been 
achieved in a few cases of streptococcal septicemia (Wright). 

Therapeutic Immunizations in Mixed Infections. 4 — "Among 
the many problems that confront the physician who would apply 



3 H. R. Varney: Jour. Amer. Med. Asso., July 27, 1907. 

4 A. P. Olmacher: Jour. Amer. Med. Asso., Vol. li, No. 7, pp, 571-572, 



THE NEPHELOMETER. 7;; 

in practice Wright's method of therapeutic bacterial immuniza- 
tion, is the one of mixed infection. In a genera] way n may 
3aid that a mixed infection oilers a less promising outlook 
than that of a single bacteria] species, hut still some brilliantly 
successful results have been obtained by inoculation in very com- 
plicated and long-standing infections. 

"(iiven two or even three bacterial species, well known ;i> 
pathogenic agents, and their simultaneous appearance in the 
secretion of a certain lesion, it is entirely proper to inoculate ;i 
mixed vaccine containing the proper closes of the offend ing bac- 
teria; or, when the urgency is not too great, inoculation with 
the predominating and most likely pathogenic agent is first to 
he performed, and in the case of an unsatisfactory issue, a vac- 
cine from the other bacterial species can be added to the subse- 
quent injections; say, for instance, that staphylococcus aureus 
or a streptococcus is found associated with bacillus coli in the 
discharge from fistulas of abdominal origin, a mixed vaccine 
would be in order. . . . 

"In dealing with mixed infections of a pyogenic nature, it 
is necessary to follow the events in the more chronic suppurations 
by bacteriologic analyses from time to time, and to modify the 
inoculations to correspond with the changes in the bacterial 
llora of the pus in case the therapeutic response is not satisfac- 
tory. . . . 

"In dealing with tuberculous diseases in which a mixed 
infection is demonstrated by the culture-test, it is proper to 
combine the inoculations of tuberculin with vaccines of one or 
more of the complicating bacterial species. . . . 

"Syphilis and a concurrent or a secondary infection with 
pyogenic bacteria, is another condition in which therapeutic 
bacterial immunization finds useful application." 

THE NEPHELOMETER.5 

With the Leishman or Wright method of estimating the 
phagocytic or the opsonic indices of the blood, it is essential to 
use bacterial suspensions containing uniform numbers of bac- 
teria in order to secure uniform results. If the suspension used 



•Jos. McFarland: Jour. Araer. Med. Asso., Oct. 5, 1907. 



74 OPSONIC METHOD. 

for one estimation contains twice as man): bacteria as that used 
for another, the results will vary according to the density of the 
suspension. The method of Wright does not provide any means 
of preparing a suspension that shall have any given number of 
bacteria, so it seemed convenient to provide some method by 
which uniform suspensions could be prepared from time to time, 
as they might be needed. 

The method suggested is that of using the density opacity 
of the suspension as an index of the number of bacteria con- 
tained, and devising some standard by which the opacity could 
be regulated. 

It was this idea that materialized in the form of the simple 
instrument to which the name nephelometer has been applied. 

It consists essentially of a series of standardizing tubes con- 
taining a suspension of a fine precipitate approximating bacterial 
suspensions in opacity and a holder for making their comparison 
easy. The standard suspensions used are precipitates of barium 
sulphate, made as follows: Two solutions, one a 13-per-cent. 
sol. of H0SO4 C. P., the other a 1 per cent. C. P. sol. BaCh, 
are prepared; then the one added to the other, so that 10 stand- 
ards, containing 99 cubic centimeters of acid and 1 cubic centi- 
meter of BaCl 2 , 98 cubic centimeters of acid and 2 cubic cen- 
timeters of BaC.2, 97 cubic centimeters of acid and 3 cubic 
centimeters of BaClo, etc., are used. 

According to the percentage or cubic centimeter of BaClo 
solution used, these tubes are denominated 1, 2, 3, 4, 5, etc. 
About 3 cubic centimeters of each solution are sealed in a small 
test-tube, care being taken to use tubes of uniform diameter, 
and also to provide similar tubes for mixing bacteria to be com- 
pared with the standard. 

Experience has shown that Standard 3 is most appropriate 
for staphylococci, and that the average phagocytosis for normal 
human blood when the corpuscles are mixed with this suspension 
and incubated for thirty minutes at 37° C, is 15 bacteria per 
polymorphonuclear. For the tubercle bacillus the most appro- 
priate density is No. 5, and the average normal phagocytosis 3 
bacteria per cell. 

If the colorless barium sulphate solution is difficult to com- 
pare with the yellowish or brownish bacterial suspensions, this 



THE NEPHELOMETER. 75 

difficulty may be overcome by placing a ground-glass screen be- 
fore the tubes and comparing the suspensions by the-use of arti- 
ficial (candle) light. 

The modus operandi of using the instrument is very simple: 
the standard-tube appropriate to the experiment is selected, 
well shaken, and placed in the left-hand pocket of the instru- 
ment, and the tube containing sterile salt solution stood in the 
other. The surface growth of the bacterium to be suspended is 
taken upon the platinum loop and stirred in, rubbing the bac- 
terial mass on the side of the tube just above the surface of the 
liquid and gradually adding the fluid to it by tilting the tube 
so as to avoid suspending masses of culture. When the mixture 
a}) pears homogeneous a comparison is made by looking through 
the instrument against the sky. If the density is insufficient 
more bacteria are added; if it be too dense, fluid can be added, 
drop by drop, from a medicine dropper, or, what is better, the 
tube can be given a few turns in the centrifuge, by which large 
clumps of bacteria are thrown down and it can be compared 
again. One should not be content with the comparison until 
the suspension be tried with others of the standard tubes than 
that which one intends to imitate. It is surprising at times 
to see how, what is thought to be an exact equivalent, turns out 
to be 1 per cent., more or less, dense. 

When the density of a ready-made vaccine is to be deter- 
mined, the procedure is varied by making the tube containing 
the vaccine the standard of comparison and changing the stand- 
ardizing tubes until the density is fully determined. 

Those who are constantly working in this field will do well 
to determine once for all the number of bacteria represented by 
each standard of density, and to make a little table for future 
use. 

A modification that is sometimes to be preferred to the fixed 
standards is that of comparing the new suspension with pre- 
viously-used suspensions, until exact uniformity is secured. The 
disadvantage of this method is that slight variations may pass 
undetected unless always compared with fixed standards, and 
that if the density of the suspensions used is not measured by 
some fixed standards, exact repetition by the experimentor him- 
self, or by those who try to repeat his work, will be impossible. 



V. 

THE BLOOD-PRESSURE. 



GENERAL CONSIDERATIONS. 

Since 1733, when Stephen Hales first succeeded in meas- 
uring the blood-pressure in the arterial system with some ac- 
curacy, until a few years ago, the greater part of experimental 
work along these lines has been done upon the lower animals, 
particularly the clog. This has been by the direct method, which 
consists in exposing a vessel, inserting a cannula, and studying 
directly the circulatory changes by observing the fluctuations of 
some form of manometer connected with the blood-vessel through 
the cannula. Obviously this method of study, while appli- 
cable for examination of the changes in the vascular system 
from a physiologic or pharmacologic standpoint, does not rep- 
resent the actual conditions existing in the human, and the data 
thus obtained can only be used to explain actual conditions in 
man by analogous reasoning. With the development of the 
indirect method of studying the circulatory phenomena, and the 
perfection of the various sphygmomanometers now obtainable, 
we are able to actually follow these changes as they occur in the 
human subject, and are able with great accuracy to note the 
effect of advancing disease, the effect of medication, and 
the intricate action of the nervous mechanism in both health 
and disease. Thus we have been able to refute or verify pre- 
vious ideas and theories as applied to the arterial system, and 
have been able to supplement and augment the knowledge gained 
from ausculatory and palpatory methods; and, finally, to have 
ante-mortem diagnoses corroborated by the post-mortem findings 
in a far larger number of cases than heretofore. 

Immediately following the practical development of the 
sphygmomanometer (Fig. 10) which is applied to the vessel in 
situ, and which affords a graphic demonstration of the pressure 
(76) 



GENERAL CONSIDERATIONS. 



( < 



and its variations in the vessel under observation, the study of 
the pulse and circulatory phenomena received a decided impetus 
which has steadily increased in strength, so that to-day a greal 
mass of corroborative and explanatory data has accumulated, 

making the study of the pulse and of blood-pressure, and the 
many factors affecting and determining them, a very important 
chapter in clinical pathology, the practical importance of which 
makes it incumbent upon every physician to study and give care- 
ful consideration to this most important subject. 




Fig. 10.— The Author's Sphygmomanometer. (Pilling & Co.) 



Blood-Pressure in its Relation to Vessel-Contraction and 
Heart-Power. — Blood-pressure is determined by several factors : 
1. The driving power of the left ventricle. 2. The condition of 
the channels through which the blood flows. 3. The condition 
of the flowing blood. In considering the question of blood- 
pressure, there must constantly be borne in mind the two prin- 
cipal factors affecting it. First, the physical laws which govern 
the flow of fluid through a system of tubes, and, second, the vital 
factor arising from the fact that the whole system is composed 
of living tissue and subject to the complicated mechanism which 
controls this. 



78 THE BLOOD-PRESSURE. 

Some of the physical factors are these : Starting from the 
left ventricle, the blood-pressure is highest in the aorta and falls 
gradually toward the capillaries. This is true of the flow of 
any fluid through a system of tubes. The fall in pressure as 
the periphery is approached, is the result of (a) friction between 
the flowing fluid and the walls of the tubes, and (b) the in- 
creased total area of the tubes on cross-section resulting from 
the repeated branching. Xaturally a thick fluid will flow less 
readily than one which is thin. 




Fig. 11.— The Author's Sphygmomanometer, showing method of 
•packing Arm-band and Bellows in case. (Pilling & Co.) 



In studying the circulation the vital factor is by far the 
most important, frequently determining the physical changes. 
The vital factor is composed of, first, the inherent tendency of 
the vessel-wall to continue in a state of partial contraction — in 
other words, to have tone, and second, the central nervous mech- 
anism controlling both the blood-vessel walls and the heart. 

It is of greatest importance to separate these two factors 
when circulatory phenomena are considered, as a careful study 
and clear knowledge of the inter-relation of the factors of both 
classes will have much to do with a proper grasp of the situation 
which is necessary to an accurate prognosis and rational treat- 
ment. 



THE SPHYGMOMANOMETER. 79 

Arterial blood-pressure is the pressure of the blood within 
the vessel under observation at the time of examination. It is 
chiefly affected by two factors. The state of the propelling mech- 
anism and the condition of the circulating blood. The first of 
these is largely dependent upon the volume of blood entering 
the arterial system at each systole, and the resistance offered to 
the escape of blood from the arteries through the arterioles and 
capillaries. 

In the clinical examination of the blood-pressure we must, 
first, constantly bear in mind the tactile sensation caused by the 
normal thickness of the vessel wall; second, the presence of 
increased tone or liypertonus producing a feeling of diminished 
diameter and increased thickness of the wall, and third, the 
occurrence of anatomic changes which may include atheroma, 
calcareous infiltration, or arteriosclerosis. 

To those desirous of following this matter under the direc- 
tion of a master mind, the little work, entitled "Arterial Hyper- 
tonias, Sclerosis and Blood-Pressure," by William Eussell, is 
highly recommended. 

THE SPHYGMOMANOMETER. 

This instrument consists essentially of two parts, (1) a 
closed pneumatic system for applying indirect pressure to a 
vessel situated in an exposed part, usually the brachial. This 
pneumatic system is so arranged that air-pressure can readily 
be applied to the arm through the medium of an inflatable rubber 
cuff connected with the measuring device, and surrounded by a 
stiff outer or protective cuff of leather or reinforced canvas; (2) 
a mercury column by which the amount of pressure existing in 
the pneumatic system is accurately indicated upon the attached 
millimeter scale. The source of pressure is a small hand-bellows. 

Method of Application. — To obtain the best clinical results 
with the sphygmomanometer it is important that all observations 
be made under as nearly invariable conditions as possible. This 
should be attained both in a series of observations on one case, as 
well as in differing cases, for the sake of more accurate compari- 
son. Attention to this detail will, in a great measure, overcome 
variations in pressure resulting from changes in position of 
different parts, etc. 



80 



THE BLOOD-PRESSURE. 



For general clinical study the arm-band (Fig. 10) should 
be adjusted to the left arm in the region of the deltoid inser- 
tion. By wrapping the rubber bag firmly about the arm and 
covering this closely with the retaining cuff, the danger of rup- 
ture of the inner cuff is avoided. In this connection a series of 
observations by the author serve to show that it is unnecessary 
to bare the arm before applying the cuff, as the interposition 
of a shirt or dress sleeve does not appreciably affect the reading. 
It is advisable, however, to remove the coat and to roll up one 



4 ■ 
a 




' 


HElHBiH ! 1 












1 2**** 




if 

__ &&*** ■:: 



Fig. 12.— The Author's Sphygmomanometer, showing Cuff applied 
and ready for blood-pressure determination. 

shirt sleeve when heavy winter underwear is worn. The tube 
emerging from the arm-band is connected with the sphygmo- 
manometer by the branch tube, which is not supplied with the 
stop-cock. The hand-bellows is connected with the branch pos- 
sessing the stop-cock; the needle-valve, situated immediately 
above this, is closed. 

The Systolic Pressure. — With these preliminaries com- 
pleted the operator finds the most accessible part of the radial 
artery at the wrist. Here the general character of the impulse 
is studied, and the vessel carefully located, with the examiner's 



BLOOD-PRESSURE DETERMINATIONS. SI 

hand in a position of case. The pulse should be counted and 
the rate recorded; then, with the pulse still under observation, 
the air-pressure in the system is quickly raised until the radial 
pulse is completely obliterated. The application of more pres- 
sure than just sufficient to accomplish this is unnecessary, and 
only adds to the patient's discomfort. The stop-cock control- 
ling the bellow's tube is then closed, and the needle-valve 
slightly opened, just sufficient to allow a gradual fall in the 
mercury column. As this fall occurs the examiner should be 
on the alert to note the return of the first impulse at the wrist. 
The height of the mercury column at this instant will exactly 
represent the measure of pressure required to obliterate the 
lumen of the artery under observation. In other words, the 
pressure in the pneumatic system approximates very closely the 
pressure exerted by the blood in the vessel under compression, 
and must, therefore, represent the pressure at its highest point 
during a heart cycle, i.e., the systolic pressure. The height of 
the mercury column at this moment is jotted down, and a sec- 
ond observation made in the same manner to corroborate the 
finding. 

The diastolic pressure may be obtained in one of two 
ways : First, after the systolic pressure has been determined, 
the fall of the mercury column is allowed to continue. As this 
occurs the mercury column will begin to show a slight oscilla- 
tory movement, which is synchronous with the pulse. At some 
point during the descent of the column of mercury, this oscilla- 
tion will reach a maximum, after which it will grow less and 
eventually cease before the column reaches zero. The base of 
the column, when the movement is at its height, represents the 
diastolic pressure. Fluctuations of the mercury corresponding 
in rhythm to the respiratory movement, or irregular variations 
due to accidental muscular movement by the patient, must not 
be confused with the pulse wave. 

In certain cases, particularly those presenting hypotension, 
it may be impossible to obtain the diastolic reading according to 
the above method. This is because the volume of the pulse wave 
or the force of th.e impulse is not sufficient to produce the neces- 
sary oscillation. In these cases the following method can usually 
be successfully employed : — 



g2 THE BLOOD-PRESSURE. 

After obliterating the radial pulse as in the examination 
for the systolic pressure, allow the pressure to slowly and steadily 
fall, at the same time carefully keep the fingers upon the pulse. 
As the pulse first reaches the wrist it will be found to be weak 
and thready, which characteristic will continue for a variable 
time, until suddenly the force of the impact will become greater 
and will assume the character of the pulse of aortic regurgita- 
tion. 

A reading of the height of the mercury column at the mo- 
ment of this change in the pulse, will indicate the diastolic 
pressure. 

The Systolic Pressure. — The systolic pressure, as indicated 
by the sphygmomanometer, represents the greatest amount of 
pressure to which the arterial wall is subjected during the con- 
traction of the left ventricle. 

The Diastolic Pressure. — The diastolic pressure represents 
the ebb to which the pressure within the vessel falls during the 
filling of the ventricles. 

The Pulse Pressure. — This figure represents the difference 
between the systolic and the diastolic pressures, and represents 
the total variation in pressure to which the vessel wall is sub- 
jected during a complete cardiac cycle. Having determined the 
systolic and the diastolic pressures in any case, it is a simple 
matter to obtain the pulse pressure by subtracting the diastolic 
from the systolic figure. 

The sphygmomanometer bearing the author's name (see 
Figs. 10, 11, and 12) has been recently devised, and embodies all 
the advantages possessed by the older models. Its individual 
points of merit are: First, the small size of the carrying case 
greatly reduces the total weight of the instrument, at the same 
time increasing its portability. Second, the simplicity of the 
mechanism and the absence of detachable parts reduces the time 
required of "setting up" to a minimum, at the same time mak- 
ing it impossible to lose or forget a part of the instrument dur- 
ing transportation. Third, the mercury employed in the mano- 
meter is never exposed, and therefore there is no opportunity 
for spilling, which might interfere with the accuracy of its 
readings. Fourth, the scale is adjustable and can instantly be 
regulated to the height of the mercury column. 



POSTURAL VARIATIONS IN BLOOD-PRESSURE. 83 

FACTORS INFLUENCING THE PULSE PRESSURE. 

It is theoretically possible that the pulse pressure should be 
increased in three ways, but it should also be remembered that 
clinically the condition of the pulse pressure bears a relation to 
all three, and that the occurrence of but one of the factors with- 
out some admixture of another, is a rarity. 

1. An increase of the amount of blood delivered by the left 
ventricle into the aorta would tend, other things being equal, to 
increase the difference between systolic and diastolic pressures. 

2. The same effect would be produced by any condition 
which would hasten the escape of blood from the vessels, inde- 
pendently of whether the blood escaped through the capillaries 
or was returned to the heart by regurgitation. 

3. Undue rigidity of the vessel walls will exert a similar 
influence upon pulse pressure, as in factor two, because the 
harder and less elastic the arteries become, the more the heart 
is compelled to move the blood throughout the arterial system 
as a whole, while the diminished elasticity would, for "mechanical 
reasons, cause the pressure during diastole to become very low. 

METHOD OF RECORDING OBSERVATION. 

For convenience in study, comparison, and for future ref- 
erence, it is advisable to formulate and adhere to some method 
of recording the blood-pressure observations. 

A chart which has been found satisfactory for this purpose, 
and which is designed to include all possible observations in this 
connection, will he found on page 281 in the Appendix. 

POSTURAL VARIATIONS IN BLOOD PRESSURE. 

Summary of observations made by 0. Z. Stephens 1 on a 
series of twenty-two healthy medical students: — 



SYSTOLIC 


STAND- 


SIT- 


SUPINE 


HEAD 


RIGHT 


LEFT 


PRESSURE. 


ING. 


TING. 




DOWN. 


LATERAL. 


LATERAL. 


Right arm . . 


.. 132.6 


133.3 


152.5 


166.2 


155.0 


110.0 


Average 


.. 130.8 


131.7 


150.4 


165.6 


143.5 


133.0 


Left arm 


. . 130.0 


130.0 


148.3 


165.0 


114.0 


156.0 


Pulse rate . . 


. . 86 


82 


68.7 


65.8 


68.1 


69.1 



1 O. Z. Stephens: Jour. Amer. Med. Asso., Oct. 1, 1804. 



84 THE BLOOD-PRESSURE. 

Summary. — 1. The blood-pressure increases in the brachials 
from the standing to the head-down posture, in the following 
order : Standing, sitting, left lateral, right lateral, supine, and 
head down. 

2. An increase in resistance is accompanied by an increase 
in heart strength. 

3. The pulse rate decreases in the same order as the blood- 
pressure increases. 

4. The decrease in pulse rate is a conservative act on the 
part of nature to protect the heart itself and the central nervous 
system. 

5. The average systolic pressure in the sitting posture was 
found to be nearly one mmHg. greater than the standing posi- 
tion. 

Physiologic Variations. — Besides the variations occurring 
in blood-pressure incident to alterations in posture, the follow- 
ing conditions have been studied in their relation to blood- 
pressure : — 

Age. — During the first year of life the systolic pressure 
usually averages from 75 to 90 mmHg. In adults it ranges 
between 100 and 130. In the aged it lies between 130 and 145. 

Excitemext usually causes temporary marked rise in pres- 
sure. 

Muscular exertion also increases systolic pressure. This 
increase becomes less marked as the individual becomes accus- 
tomed to performing the act or acts. The gradual reduction 
following systematic exercise is one of the benefits of training. 

Sleep. — In the early hours of sleep there is quite a fall 
in blood-pressure, which gradually rises toward morning (Brush 
and Fairweather). 

Pathologic Temporary Increase in Blood-Pressure. — Drugs 
may increase the blood-pressure either by acting upon the heart 
or by causing contraction of the peripheral circulation. Acute 
asphyxia or acute anemia of the brain will stimulate the vaso- 
motor centers, causing a marked rise in blood-pressure. 

Pain, even when slight or momentary, will cause a rise in 
blood-pressure even in normal individuals. 

Pathologic Temporary Diminution in Blood-Pressure. — 
Widespread dilatation of the blood-vessels from any cause may 



THE CLINICAL VALUE OF THE SPHYGMOMANOMETER. 85 

cause serious diminution in blood-pressure. Lack of splanchnic 

tone and wide dilatation of the abdominal vessels may have a 
fatal termination mainly through the great diminution in blood- 
pressure occasioned, the patient literally bleeding to death in 
his own abdominal vessels. A somewhat similar condition exists 
in chloral and alcohol poisoning, also in collapse. 

Crile's exhaustive experiments seem to show that the con- 
dition known as surgical shock is caused by an exhaustion or 
paralysis of the vasomotor center, which renders it incapable of 
maintaining the normal vascular tone. 

THE CLINICAL VALUE OF THE SPHYGMOMANOMETER. 

The sphygmomanometer is especially applicable to the 
problems of medicine involving cardiac and renal disease, and 
should be constantly employed to furnish an index of the gravity 
of the condition, and as an aid in therapeutics by furnishing 
a graphic record of the effect of medication upon the cardio- 
vascular system. Xo pathologic condition of either the heart or 
the kidneys, or of the two in combination, can be thoroughly 
understood or rationally treated without a comparative study of 
the state of the circulation, blood-pressure, etc., and, without 
such a knowledge, the management of the case must of necessity 
be uncertain because of the element of guesswork, which can be 
eliminated in no other way but by employing this most valuable 
aid. 

The great amount of painstaking work that has been clone 
by many investigators along these lines, has resulted in the 
development of many clearly recognized and carefully classified 
clinical pictures, a knowledge of which will often lead in the 
right direction by shedding much light upon the inter-relation 
of apparently dissociated facts. 

In Cardiac Disease. — The sphygmomanometer finding alone 
may be sufficient to determine the diagnosis. In aortic regurgi- 
tation the large volume of blood projected into the aorta by each 
systole must of necessity produce a high systolic reading, while 
subsequently, owing to the disposal of blood in two directions, 
through the capillaries and back into the left ventricle, the 
diastolic fall will be great. The combined result of these two 



86 THE BLOOD-PRESSURE. 

extremes will produce an enormous pulse pressure, this differ- 
ence becoming even greater in the presence of markedly sclerotic 
and hardened arteries. 

Therapeutically, we are enabled more accurately to follow 
the action of the drugs employed. Thus we obtain the fullest 
effect from the measures available, without overtreating, and are 
at the same time guarded against the administration of improper 
or useless remedies. 

In the Diagnosis of Heart-Muscle Degeneration. — The 
sphygmomanometer is frequently the earliest means of detect- 
ing diminished heart-power from beginning degeneration of the 
muscle, this evidence being readily obtainable long before the 
development of the usual physical signs. This condition may 
be detected in the following manner: After applying the in- 
strument, while palpating the pulse at the time when the pres- 
sure is gradually being raised, it will occasionally be noted that 
just before obliteration of the pulse some systoles fail to reach 
the wrist, showing undoubted irregularity in the power of the 
"heart's action, the weaker impulses being prevented by the ob- 
struction from reaching the wrist. This phenomenon can be 
very readily demonstrated in a pulse which to ordinary palpatory 
methods is apparently regular in force and rhythm. 

In Renal Disease. — Here we may be enabled to confirm the 
diagnosis of incipient or latent Bright's disease by a study of 
the blood-pressure. In the presence of a slight recurrent albu- 
minuria with beginning arteriosclerotic change and a systolic 
pressure which remains persistently above the normal in spite of 
appropriate treatment directed toward the hypertension, the 
probable diagnosis would be that of beginning renal change. 
Thus by an early warning we may be enabled to check the ad- 
vance of serious symptoms or at least to considerably prolong the 
patient's period of activity and usefulness. 

In cases of both acute and chronic lesions of the kidneys, it 
will be almost uniformly noted that the blood-pressure, as indi- 
cated by the sphygmomanometer, is permanently increased. In 
chronic cases the systolic not infrequently reaches 200 or 225 
mmHg., while occasionally it may be above 250. 

Friecllander 2 states that if this high pressure lasts for more 
than four weeks, hypertrophy of the heart is sure to ensue. 

2 Friedlander: Arch. f. Physiol., 190L 



THE CLINICAL VALUE OF THE SPHYGMOMANOMETER. 87 

In primary acute Bright's disease and in nephritis follow- 
ing scarlet fever, there is probably always an increase in arterial 
pressure. Sudden and marked rise in pressure has been noted 
within forty-eight hours of the onset of an acute nephritis. 3 

In advanced nephritis the study and detection of high ten- 
sion and hard pulse will indicate the urgent necessity for ener- 
getic treatment for the reduction of blood-pressure; thus the 
brain may be saved from the serious effects of continued high 
pressure, and the patient protected from the occurrence of coma 
or apoplectic attacks. 

In angina pectoris the attacks may be lessened in number 
and severity by timely detection of the increased tension result- 
ing from arterial spasm, the effect of diet, and medication can 
be followed; and as have been reported, under favorable con- 
ditions, a few patients will be, to all intents and purposes, 
cured. 

Diabetes: Its Relation to Cardiac Complications. 4 — In a 
series of 25 cases, all ages, rising both Stantom's and the Eiva- 
Eocci apparatus; 12 and 18 centimeter arm-bands; 150 ob- 
servations. Diastolic pressure hard to obtain. 

Summary of Systolic Pressure Records. 

Number of cases 25 

Average age 45 years. 

Male 13 

Female 12 

Average weight 156 pounds. 

Average systolic blood-pressure 127 mmHg. 

Average of cases showing 3 per cent, sugar 121 mmHg. 

Showing less than 3 per cent, sugar, pressure average .... 135 mmHg. 

Xumber of cases developing acid intoxication 10 

Average systolic pressure 107 mmHg. 

Xumber of cases not showing acid intoxication 15 

Average systolic pressure 140 mmHg. 

Number of cases showing indication of arterio-sclerosis 

and kidney involvement 5 

Average systolic pressure 164 mmHg. 

Exophthalmic Goitre. 5 — The systolic pressure varies, some- 
times low, usually higher than normal, occasionally very high. 
The pulse pressure may be very great, showing probably large 
systolic output from the left ventricle. 

3 Buttennann: Arch. f. Klin. Med., Vol. lxxiv, p. 1. 

4 Arthur B. Elliott: Jour. Amer. Med. Asso., July 6. 1907. 

5 Reported by L. F. Baker: Jour. Amer. Med. Asso., Oct. 12, 1907. 



88 THE BLOOD-PRESSURE. 

Lead Colic and Beginning Peritonitis. — These two condi- 
tions are usually associated with high arterial pressure. 

Scarlet Fever. — The blood-pressure curve follows ver) T 
closely the pulse and temperature curve. After the seventh or 
eighth day the blood-pressure is usually below normal. Com- 
plications have a marked effect upon blood-pressure. Signs of 
increased arterial tension are present in most, if not all, cases 
which show albuminuria; the high pressure is usually syn- 
chronous with the onset of the albuminuria. This is always 
accompanied by a marked slowing of the heart's action. With 
the subsidence of the nephritis the pulse-rate increases in the 
ratio that the arterial pressure diminishes. 6 

Diphtheria. — A reduction in blood-pressure will be found 
in every case, depending to a large extent, if not entirety, upon 
the degree of toxemia. The administration of strychnia and 
alcohol annul the fall of pressure, but only if given regularly. 

The administration of antitoxin, while causing a slight rise 
in temperature, has no effect upon the blood-pressure (J. David- 
son). 

Typhoid Fever. — Here the blood-pressure is always consid- 
erably lowered, the minimum depressing steadily the longer the 
duration of the case. The pressure rises gradually to normal 
with the establishment of convalescence or through the inter- 
occurrence of a complication, as pneumonia or peritonitis (J. 
Davidson). 

THE EFFECT OF BLOOD-PRESSURE ON THE 
FLOW OF URINE. 

The amount of urine passed varies directly with the pulse 
pressure. 7 High arterial pressure, by sending more blood to the 
kidneys, increases the flow of urine. 

Lowered general arterial pressure will cause a diminution 
in the quantity of urine; this diminution is augmented if for 
any reason there be also an increase in venous pressure. 



3 J. Davidson : Lancet, Oct. 19, 1907. 

' Erlanger and Hooker: Amer. Jour. Physiol., Vol. x, p. 7. 



VI. 

COAGULATION TIME. 



GENERAL CONSIDERATIONS. 

The results obtained by improved clinical methods have 
only . recently been reduced to a sufficiently accurate basis to 
admit of much confidence being placed in them, or to permit 
of their use in clinical medicine. 

The time required for coagulation of the blood depends 
upon many conditions. This fact makes attention to detail and 
uniformity . of technic of the utmost importance. 

The chief factors influencing the coagulation-time are: 1. 
The time during which the shed blood remains in contact with 
the tissues, coagulation being slower from a deep than from a 
superficial incision, the difference in time may amount to as 
much as three minutes (Emerson). 2. The amount of pressure 
applied to the tissues to start the flow. 3. Upon the amount of 
blood shed. 4. Upon the nature of the vessel receiving the blood 
and its temperature. Factors of less importance are : time of 
day, atmospheric humidity, relation of time of shedding to diges- 
tion, ingestion of drugs, etc. 

THE SPECIMEN. 

This should be taken midway between meals, and not 
after the ingestion of drugs, which may influence coagulation- 
time. The tip of the finger or the lobe of the ear should be 
thoroughly cleansed and dried, and a sufficiently large puncture 
made with a flat-bladed instrument (Daland lancet) to insure 
free and sufficient flow. 

METHODS OF DETERMINATION. 

Milan's Method. — Place a drop of blood upon the center of a 
large glass slide or watch crystal. After a minute or two gently 

(89) 



go COAGULATION TIME. 

tilt the glass from side to side. This is continued at short in- 
tervals until the drop of blood, which at first assumed a pear 
shape when the glass was tilted, is seen to assume the form of 
a blunt and symmetrical cone. This change in the character 
of the drop indicates the completion of coagulation. The aver- 
age time of coagulation of normal blood by this method is about 
five minutes. 

This method is at best imperfect and uncertain, since none 
of the modifying factors are controlled. 

Wright's Method. — This is a far more accurate method, 
since in a large measure it controls and renders uniform the 
majority of the variable factors. 

Wrights coagulometer consists of the following parts 
(Fig. 13) :- 

Copper reservoir with numbered grid and rubber mat. 

Pinchcock and tube. 

Thermometer marked 18.5° and 37° C. 

Twelve pipettes numbered 1 to 12. 

Wire for cleaning tubes. 

Lancets in bottle. 

Aspirating tube. 

Bottles for ether and alcohol. 

The Principle of the Method. — A series of tubes of stand- 
ard caliber filled with a definite amount of blood. The coagula- 
tion is observed at a standard temperature, and the coagulation- 
time is determined by blowing the contents of each tube upon 
a piece of clean white blotter or filter paper, at increasing in- 
tervals from the time of filling. 

The tubes are 10 centimeters in length and have an internal 
caliber of 0.25 millimeters; they are marked at a point 5 cen- 
timeters from the lower end of the tube (Fig. 1). The coagula- 
tion-time is usually determined at 18.5° C. This temperature 
has been chosen as a standard because it approximates the tem- 
perature of the hospital ward or bed-room in temperate climates, 
and because this temperature delays the coagulation sufficiently 
long to allow the coagulation-time being, even in cases of rapidly 
coagulating blood, determined with considerable accuracy. 

In case of slowly coagulating blood it is convenient to carry 
out the determination at 37° C. (blood-heat). 



METHODS OF DETERMINATION. 



91 



The tubes are to brought to a standard temperature before 
they are filled with blood. For this purpose close-fitting rubber 
caps are put on both ends of the coagulation-tubes, which are 
then placed butt end downward in the reservoir containing water 
of the standard (18.5° C.) temperature (see Fig. 2 A). 

Fig.2. 




Fig. 13.— Wright's Coagulometer. 1. Capillary Tube. 2. Copper 

Reservoir. A. Capillary Tube cooling. B. Tube filled 

with blood. C. Thermometer. (Hawksley.) 

After sufficient time has elapsed to allow the tubes to ac- 
quire the desired temperature, they are removed from the water 
and the rubber caps removed. Great care should be exercised to 
prevent moisture from entering the bore of the tubes. 



92 COAGULATION TIME. 

Method of Filling the Tubes. — The blood which flows from 
the puncture is drawn into the tube with the least possible delay. 
A column of blood approximately 5 centimeters in length, is 
drawn into each tube, and then this column drawn a few centi- 
meters from the tip of the tube. No important fallacy will arise 
if the column of blood be slightly longer or shorter than the 
5 centimeters. 

Before beginning the observation it is well to have the tubes 
arranged in the order in which they are numbered. In ordinary 
cases from six to eight tubes are employed. For cases where 
coagulability is greatly reduced, a larger number of tubes will 
be required, unless the coagulation-time has been approximately 
determined by a preliminary test. 

A watch provided with a second-hand, and paper and pencil 
for noting the time of filling and blowing out, should be at hand. 
If an assistant is available, the task of keeping record of the time 
may be conveniently committed to him. 

It is of decided advantage, before commencing, to set the 
second-hand so that it coincides with the minute markings 
around the dial. As each drop of blood is drawn, the time (min- 
utes and seconds) is recorded. 

It will be found convenient to allow an interval of ten or 
fifteen seconds between the filling of successive tubes. 

As each tube is filled it is returned to the tank. It is not 
necessary to replace the rubber caps, as the air-space at the top 
of the tube is sufficient to prevent the water coming in contact 
with the blood in the tube. The viscosity of the blood will be 
sufficient to prevent the blood being forced further up the tube. 

During the time that the tubes remain in the reservoir the 
temperature of the water must be maintained by the addition 
of hot or cold water from time to time as necessary. 

After the lapse of exactly two minutes from the filling of 
tube No. 1, it is removed from the water and its contents blown 
upon a piece of white blotting or filter paper. If the contents 
cannot be blown out coagulation is noted down as complete. If 
the contents can be blown out, but shreds of fibrin are seen 
adhering to the interior of the tube, coagulation is recorded as 
incomplete. If the contents can be blown out cleanly and im- 
mediately sink into the paper, coagulation has not yet begun. 



METHODS OF DETERMINATION. 



9^ 



If in the first tube (two minutes) coagulation has been 

found to be complete, the second tube is removed at the expira- 
tion of one and three-quarter minutes, and the contents treated 
in the same manner. If this also is found clotted, the next in 
the Beries is tested at a still shorter interval, and so on until a 
tube is found in which coagulation is incomplete. 

If the blood in the first tube, after the expiration of two 
minutes shows coagulation to be incomplete, a slightly longer 
interval is allowed before the second tube is blown out, and so 
on until one is found which shows coagulation to be complete. 

The same process is carried out if, in the first tube coagu- 





Fig. 14.— Bogg's Coagulometer, diagrammatic representation. 



lation has not yet begun, except that a still longer interval is 
allowed to pass before the second tube is blown out. 

Permissible Margin of Error. — If the directions wdrich 
have been outlined above are adhered to, the margin of error 
in the coagulation-time should not be more than five seconds, 
and should never exceed fifteen seconds. 

The normal coagulation-time by this method lies between 
three and five minutes. 

Method of Russell and Brodie. — This method requires a 
microscope. The coagulometer consists of a small, moist cham- 
ber with a glass bottom (Fig. 14:), which can be placed upon the 
stage of the microscope. This is fitted with a truncated cone of 
glass which projects downward into the moist chamber. The 
end surface of this cone is of definite size (about 5 millimeters 



94 COAGULATION TIME. 

diameter), and on it is placed the drop of blood, care being 
taken to see that the drop of blood only just covers the surface, 
hence is always of the same size. The cone is then quickly fitted 
into the moist chamber to prevent alterations due to drying, 
temperature, etc. Through the side of the moist chamber pro- 
jects a fine tube, through which, by means of a hand-bulb, a 
gentle stream of air can be projected against the blood. The 
preparation of the slide being completed, the drop of blood is 
observed with the low power of the microscope. 

The blowing should be done at as long intervals as possible, 
and also very gently. The corpuscles will first be seen to move 
freely as individuals ; later, as coagulation begins, the corpuscles 
will no longer move freely in the drop, but the drop will begin 
to change shape en masse. Finally, there will occur only elastic 
motion, and the part of the drop displaced by the current of air 
will spring back to its original position as soon as the current 
of air ceases. This is to be taken as the terminal point, as now 
only can the clot be demonstrated, by quickly removing the slide 
and touching the drop to a piece of dry filter paper. 

This apparatus is probably the best method yet devised for 
the determination of coagulation-time. Eecently this instrument 
has been improved upon by Boggs, who has substituted a metal 
tube and an improved cone. 

With the improved instrument Emerson (clinical diagno- 
sis) reports normal variations in the coagulation-time between 
three and eight minutes, with an average of five minutes, eight 
seconds. 

CLINICAL OBSERVATIONS. 

The following results were obtained by Drs. Eobertson, Ill- 
man, and Duncan with the Wright apparatus 1 : — 

The normal clotting time was found to be from two minutes, 
thirty seconds, to two minutes, forty-five seconds, using half 
blood-heat (18.5°C). 

Typhoid Fever. — In the acute febrile stage the average 
clotting time obtained from observations on 61 cases was three 
and one-third minutes. A lengthened clotting time was found 



1 Report of Section on Pharmacology and Therapeutics, Amer. Med. Asso., page 

51, et seq., 1907. 



CLINICAL OBSERVATIONS IN DISEASE. 95 

to indicate impending hemorrhage. If, after hemorrhage, the 
usual shortening of the clotting time does not occur, then an- 
other hemorrhage is imminent. This shortening of coagulation- 
time is also noted after hemorrhage accompanying abortion and 
carcinoma of the uterus. 

In typhoid fever the clotting time returns gradually to nor- 
mal, as the temperature declines with the establishment of con- 
valescence. 

In cases showing abnormal clotting time, the administration 
of the many drugs supposed to favor this condition, both sepa- 
rately and combined, were found to have no appreciable effect 
upon the condition. 

The following averages were obtained from observations on 
a large number of patients : — 

Specific meningitis 3.10 

Coal gas poisoning 4.05 

Gastric ulcer 4.10 

Salpingitis 5.15 

Alcoholic gastritis 3.10 

Apoplexy 3.05 

Carcinoma « 4.35 

Jaundice 6.30 to 8.30 

Rheumatism 3.00 to 4.00 

Pneumonia 2.47 

Xephritis 2.25 

Diabetes . . . . 2.55 

Exophthalmic goitre 2.45 



VII. 

BLOOD PARASITES. 



THE PLASMODIUM MALARIA. 

Three forms or varieties of these organisms are recognized : 
the tertian, quartan, and estivo-autumnal. The cycle of growth 
of each of these differs. As the characteristic organism of the 
group, the plasmodium of tertian fever alone will be described, 
together with a brief differential table. 

The Tertian Variety. — In the early stage this is a small, 
hyaline body with somewhat indistinct outlines occupying only 
a small portion of the interior of the red blood-cell, and usually 
situated eccentrically. It is living, and exhibits rapid and dis- 
tinct ameboid movements. The hyaline body rapidly increases 
in size. Small groups of actively moving, pigment granules 
appear about this body, and the surrounding red cell becomes 
pale and swollen. When the parasite has attained its full 
growth, the outline of the red cell is hardly distinguishable. 
The activity of the ameboid movements gradually diminishes, 
and the pigment granules arrange themselves around the cir- 
cumference of the cell. The final stage of development then 
begins. Signs of segmentation appear around the periphery 
of the organism, and the pigment granules pass toward and 
collect in the center. Finally, when segmentation is complete 
a "rosette" is formed by the symmetrical arrangement of the 
segments, sixteen to twenty in number radiating from the cen- 
tral mass of pigment. The tertian parasite passes through this 
cycle in forty-eight hours. 

Quartan Variety. — The preceding description answers, ex- 
cept that the organism is smaller, the pigment less active and 
coarser. The segments number five to twelve, and the cycle of 
development is complete in seventy-two hours. 

Estivo-Autumnal Parasite. — This organism also begins as 
(96) 



THE DETECTION' OF THE PLASMODIUM 



\r t 



a hyaline body and develops in a similar manner. The parasites 
are smaller, the pigmenl granules scant and motionless. Seg- 
mentation occurs in the internal organs only, so that these forms 

are never found in blood obtained from the peripheral circula- 
tion. The organism requires twenty-four to forty-eight hours 
for its cycle. After the disease has lasted a week or more, pig- 
mented crescents and spherical or oval bodies are found, which 
appear to be pathognomonic of this variety of parasite. 

Apart from the usual life cycle, each form is capable of 
giving rise to extracellular, vacuolated, and flagellate bodies. 



TERTIAN. 

Develops in 48 hours. 

Pale and indistinct. 

Actively ameboid. 
Pigment fine. 

Pigment actively mo- 
tile. 
Pigment light. 
Full-size of corpuscle. 

Degeneration forms 
twice the size of cor- 
puscle. 

Segments 16 to 20. 

Irregular eegmcnts 
are frequent. 

Corpuscles large, 
colorless and swol- 
len. 



QUARTAN. 

Develops in 72 hours. 

Sharp outline, re- 
fracts. 
Slightly ameboid. 
Pigment coarse. 

Pigment slow in move- 
ment. 

Pigment dark. 

Smaller than corpus- 
cle. 

Degeneration forms 
vary, much smaller 
than Tertian. 

Segments 5 to 12. 

Beautiful rosettes. 

Corpuscles shrunken 
and brassv. 



ESTIVO- AUTUMNAL 

Develops in 24 to 48 
hours. 

Has irregular appear- 
ance. 

Actively ameboid. 

Pigment granules very 
few. 

Pigment still. 

Pigment light. 
Smaller than corpus- 
cle. 
Absent. 



Segmentation forms 
not seen in peri- 
pheral blood. 

Corpuscles shriveled, 
brassy, not decolor- 
ized. 

Forms crescents. 



It is evident that while there is considerable similarity 
among the three varieties of parasites, there are, nevertheless, 
certain peculiarities and characteristics which enable the expert 
to quite positively differentiate between them. 



THE DETECTION OF THE PLASMODIUM. 

In searching for the organism it is always desirable, when 
possible, to examine the fresh specimen at the bedside of the 
patient. If this is impracticable, warm the slide and seal the 
cover with a little vaseline, and the organisms wall retain their 
motility for at least a couple of hours. 



98 BLOOD PARASITES. 

If considerable time must elapse before the examination, 
films should be made, dried and stained, by one of the poly- 
chrome (Eomanowski) stains. 

Cautions. — Do not have the blood-films too thick; the 
individual cells should not overlap; there should be no rouleau 
formation. Do not give up the search until all the films have 
been examined thoroughly. A number of fields may show no 
organisms, then some may appear. The specimen is best ob- 
tained either eight or ten hours before or after a chill, as at this 
time the organisms are most likely to be in the peripheral cir- 
culation. Medication should be withheld for as long a time 
before the specimen is taken as possible. 

What to Examine For. — Fresh specimen: Look for red 
cells containing actively moving black specks (pigment granules 
and living protoplasm). Unusually pale cells containing clear 
areas which are irregular and constantly changing shape. Extra 
large red blood-cells. 

After a little experience the pigmented organisms are read- 
ily distinguished. Violent commotion among a group of red 
cells will direct attention to the flagellate. The hyaline bodies 
are usually the most difficult to identify without experience. 
The apparent, but not real (artefact), may be found in large 
numbers; these are usually distorted and deformed cells that 
have become altered in the preparation of the specimen. Finally, 
pigment-bearing leucocytes may be found in excess in malaria. 
These are, as a rule, polymorphonuclears, which have become 
phagocytic and have taken up the iron pigment, set free by the 
malarial organisms. 

THE CLINICAL RELATIONS OF THE PLASMODIUM. 

Segmentation of the organisms immediately precedes the 
chill and fever, consequently the segmentation forms will be 
present in the peripheral circulation at the beginning and at 
the height of the paroxysm, while the hyaline forms are discov- 
erable during and after the seizure. 

As the tertian parasite requires forty-eight hours to develop, 
segmentation and parasites will occur every other day, provided 
that only one group or brood of parasites be present. More com- 
monly two groups of different ages exist (double tertian), seg- 



FILAEIA SANGUINIS HOMINIS. 99 

mentating on alternate days and producing the quotidian (daily) 
type of fever. 

The quartan parasite, requiring seventy-two days to mature, 
gives rise to a paroxysm with a two-day interval of ease. If 
there are two broods there will occur two days of paroxysm, fol- 
lowed by one day of rest (double quartan), while three groups 
will cause a daily or quotidian attack like the double tertian. In 
these multiple infections the time of fever need not occur at 
the same hour on successive days. 

The estivo-autumnal type has variable periods of develop- 
ment. Segmentation and fever may be very irregular, and in 
some cases continuous. 



SPIROCH/ETE OF RELAPSING FEVER. 

The specific organism of relapsing fever may be found in 
the fresh specimen of blood. Prepare this in the same manner 
as for the malarial parasite. This organism appears as narrow 
spiral filaments from 36 to 40 microns in length. They are 
actively motile, and attract attention by the active movement 
imparted to the red blood-cells in their vicinity. They are in 
the peripheral blood only during the fever, when they appear 
in large numbers. Many may be seen in a single field of the 
microscope. 

FILARIA SANGUINIS HOMINIS. 

There are several forms of this parasite ; for special distin- 
guishing characteristics, larger works must be consulted. (See 
also page 113.) The most important of the group is the filaria 
nocturna, which is supposed to be responsible for certain forms 
of chyluria, elephantiasis, and lymph-scrotum. The adult or 
parent organism is slender and thread-like, varying from three 
to six inches in length. It inhabits the lymphatics and tissues. 
The embryos average in length about one-seventy-fifth of an inch, 
and are about as wide as a red blood-cell. The search for the 
embryos should be made both in the day and the night. Study 
under low power a fresh drop of blood. These embryos are 
decidedly active, and create a commotion among the neighboring 
cells. 



100 BLOOD PARASITES. 

TRYPANOSOMIASIS. 

This is the organism of the so-called sleeping sickness. 
There are a large number of varieties of this organism which 
infect man and the lower animals. The trypanosoma gambiense 
can be demonstrated in the majority of cases. It is described 
as varying in length from 20 to 25 microns, by about 2 to 2.5 
microns in width. It is distinguished by a single flagellum 
which extends through almost the entire length of the organism, 
and extends beyond its anterior end. When alive it has a slow, 
spiral undulating movement. (See page 106.) 



LEISHMAN-DONOVAN BODIES. 

These bodies were first found in the spleen of patients who 
were apparently suffering from an irregular form of malarial 
fever. The body is small or elliptical, from 2 to 3 microns 
in diameter, containing chromatin. The bodies occur either 
free in the plasma or imbedded with others in a matrix or 
zooglia, often as many as a dozen being in one cluster. They 
are believed not to occupy the bodies of the red blood-cells. They 
are not demonstrable in the peripheral blood. 

Staining. — Wright's modification of the Eomanowsky 
stain : the chromatin dark, the cell-body blue, and the zooglia 
a fainter mauve. 



VIII. 

ANIMAL PARASITES. 



THE ANIMAL PARASITES OF MAN. 

By "animal parasites of man" is meant animal organisms 
which live temporarily or permanently on or in the human body, 
and which receive their nourishment therefrom. 

Temporary parasites include the ectoparasites (epizoa). 
These may inhabit the skin, conjunctival sac, the mouth, the 
nose, and the accessory sinuses. A familiar example of this 
class is the sarcoptes scabiei or itch mite. 

Most of the permanent or stationary parasites are found in 
the internal organs, and belong to the class of entoparasites or 
entozoa. Many of these parasites, such as the teniae, ascarides, 
and ankvlostoma, inhabit man only when mature; others, of 
which the echinococcus is an example, inhabit man only during 
a certain period of their existence. Thus man may be either 
the actual or only the intermediate host. Again, for many para- 
sites, such as the tenia solium and the tenia saginata, man is 
the only host. Finally, man may also become the host for para- 
sites which, as a rule, select some other animal. Thus the cysti- 
cercus cellulosse, common to the pig and the cat, may occasion- 
ally be found in man. 

In this section the classification of Max Braun 1 has been 
adopted, and the majority of the descriptions of the parasites 
which follow have been abstracted from this w r ork. 

Xo attempt has been made to include the rarer forms of 
parasites, as these are considered to be beyond the scope of this 
work. 



1 English Translation of "The Animal Parasites, of Man." (Wood & Co., 1906.) 

(101) 



102 ANIMAL PARASITES. 

CLASSIFICATION OF THE MORE COMMON ANIMAL 
PARASITES OF MAN, 

A. Protozoa. 

Class I. Ehizopodia. — Ameba coli (Loesch). 

Class II. Flagellata (Mastigophora). 

(a) Trichomonas: 

1. Trichomonas vaginalis. 

2. Trichomonas intestinalis. 

3. Trichomonas pnlmonalis. 

(b) Circomonades: 

1. Lamblia intestinalis. 

2. Tripanosoma. 

Class III. Sporozoa. 
Coccidia: 

1. Coccidium perforans or hominis. 

2. Hemosporidia. 

Class IV. Ixfusoria. — Balantidium coli or Parame- 
cium coli. 

B. Platyhelminthes (Flat worms). 

Class I. Trematoda (End). 

1. Fasciola hepaticum syn. distomum hepaticum. 

2. Distomum pulmonale syn. distomum vestermani. 

3. Distomum lanceolatum syn. dicrocelium lanceo- 

latum. 

4. Distomum hematobium syn. bilharzia, syn. schis- 

tosomum hematobium. 

Class II. (Bud.) 

(a) Botliriocephaloidia : 

1. Bothriocephalus latus. syn. tenia lata. 

(b) Teniidce: 

1. Tenia nana. 

2. Tenia lanceolata. 

3. Tenia solium. 

4. Cysticercus acanthotrias. 

5. Tenia saginata or mediocanellata. 

6. Tenia echinococcus. 



PROTOZOA. 103 

C. Nematoda (Thread worms). 

1. Strongyloides (rhabdonema strongyloides) syn. anguil- 

lula intestinalis et stercoralis. 

2. Filaria sanguinis hominis. 

3. Triehocephalus dispar (whip worm). 

4. Trichina spiralis. 

5. Ankylostoma duodenale. 

6. TJncinaria Americana. 

7. Ascaris lumbrieoides. 

8. Oxvuris vermicularis. 



A. PROTOZOA. 

The protozoa is a microscopic living organism. It is mono- 
cellular and represents the lowest form of animal life. The 
substance of the body consists of a finely granular, contractile 
protoplasm, which may be mono- or poly- nuclear. The viscid 
hyaline entosarc is capable of motion by expansion and con- 
traction, or by the extension and retraction of pseudopodia, 
cilia or flagella. Propagation takes place by segmentation or 
gemmination. 

CLASS I. RHIZOPODIA. 

The ameba coli produces the well-known amebic dysentery. 
The ameba in man is, however, not confined to the intestines, 
but has been found in the pus of liver abscesses, in pleuritic and 
peritoneal exudates, in the mucous membranes, and in tumors 
of the urinary bladder. 

Characteristics. — This organism is an ameboid body meas- 
uring from 20 to 30 microns in diameter. It is composed struc- 
turally of a clear protoplasmic outer portion, ectosarc, and a 
finely or coarsely granular central portion, entosarc, which usu- 
ally shows a number of clear vacuoles and one or more nuclei 
(Plate I, a). When living it shows active ameboid movements 
which are greatly increased if the organism is kept warm. In 
the living state the cell frequently includes foreign bodies, such 
as bacteria, pigment granules, and fragments of blood-corpuscles 
and other cells. 



104 ANIMAL PARASITES. 

Locomotion is accomplished by irregular extension and re- 
traction of pseudopods, which are thrown out from the periphery. 
These pseudopods are at first composed of the clear outer por- 
tion, which, as the projection gradually increases, includes the 
central granular zone. When surrounded by unfavorable envi- 
ronment, the organism undergoes a form of change known as 
the encysted state. In this state the body becomes spherical, 
and the outer wall thickened, while the division of the cell into 
two portions is lost, the whole becoming uniformly granular. 

Method of Examination. — The fecal discharges should be 
caught in a warm receptacle and immediately transferred to the 
laboratory for examination. If it is necessary to keep the 
specimen for a short time, this may be accomplished by imme- 
diately placing the specimen in a thermostat at body tempera- 
ture, where it should remain until transferred to the warm stage 
of the microscope. Preservation of specimens for more than a 
few hours is unsatisfactory, because even under the favorable 
circumstances of heat and moisture obtaining in the thermostat, 
motility is rapidly lost, and at the expiration of twenty-four 
or thirty-six hours the organisms are no longer discoverable. 

The Warm Stage. — A convenient method of maintaining 
a warm stage for examination and study of this organism, is as 
follows : A flat strip of copper or of brass, three inches wide 
by six or eight inches long, is perforated by a half or three- 
quarter inch aperture, situated in the center of one end at a 
distance of about one inch from the free maroin. The stao-e 
of the microscope should be covered with several thicknesses of 
asbestos paper or felt, upon which the metal sheet is clamped 
so that the openings in the stage and in the copper-strip coin- 
cide. A Bunsen burner or alcohol lamp is adjusted under the 
outer extremity of the strip, so that when the metal has attained 
its maximum heat the portion immediately surrounding the 
aperture is maintained at about body-temperature. A specimen 
placed upon a slide, in position for examination, will maintain 
the motility of the ameba for a number of hours. 

It will be found convenient to examine the spread, first by 
the low power, then, after locating a suitable portion of the 
slide, the higher objective may be swung in and the organisms 
studied in detail. The important aids to successful search are 



PLATE I. 




Intestinal Parasites of Man. (Redrawn from Max Braun) 

a, Amebae coli. b, Trichomonas vaginalis, c, Trichomonas intestinalis. 
d, Lamblia intestinalis. e, Tripanosomes. /*, Coccidium hominis. 



PROTOZOA. 105 

a thin and uniform spread of Liquid feces, and a low, slightly 
oblique illumination. 



CLASS II. FLAGELLATA. 
a. Trichomonides. 

1. Trichomonas Vaginalis (see Plate 1, b). — The form 
of the body is very variable, elongated, fusiform or bean-shaped. 
It is ameboid. The length varies between .015 to .025 milli- 
meters in length, by .007 to .012 in breadth. The posterior ex- 
tremity is drawn out into a point and is about one-half the 
length of the remainder of the body. The cuticle is very thin 
and the body-substance finely granular. At the anterior ex- 
tremity there are three or four flagella which are of equal length, 
and which are firmly united at their base, but which easily fall 
off. There is an undulating membrane which moves spirally 
across the body, arising from the base of insertion of the flagella 
and terminating at the base of the caudal process. The nucleus 
is vesicular, elongated, and situated in the anterior extremity. 
Propagation occurs by division. 

2. Trichomonas Intestinalis (see Plate I, c). — Some 
authors believe this organism to be identical with the trichomonas 
vaginalis. It is described as being .01 to .015 millimeter in 
length, the posterior extremity terminates in a point with a 
row of cilia. These commence at the anterior extremity and 
extend over the body. 

This organism has been found in the urethra of man, the 
vagina, the large and small intestines of both healthy and sick 
persons, in the stomach and in the oral cavity. 

3. Trichomonas Pulmonale. In all probability this is 
identical with the preceding. 

b. Cercomonides. 

1. Cercomonas Intestinalis or Lamblia Intestinalis 
(see Plate I, d). — Length, .01 to .020 millimeter, and width, 
.005 to .012. The flagella are of about equal length (.009 to 
.Ol-t millimeter). The body is finely granular, and has a very 
thin cuticle which does not entirely prevent changes in the form 
of the body. The very motile tail-like appendages in the frontal 



106 ANIMAL PARASITES. 

plane are flattened; the excavation at the anterior extremity 
which is directed obliquely forward and with its border project- 
ing backward, no doubt corresponds to the peristome, and serves 
as a clinging organ. The first pair of flagella (anterior flagella) 
arise from the anterior edge of the peristome; the second and 
third pairs (lateral and median) from the posterior edge, whereas 
the tail flagella are inserted at the posterior end of the tail. In 
life the anterior and lateral flagella sway only with the part pro- 
jecting from the body; the median and tail flagella are quite 
free. The anterior flagella appear to be connected with the nu- 
cleus. The nucleus is dumb-bell shaped and has a nucleolus in 
each half, and lies anteriorly in the part of the body which is ex- 
cavated. This organism has encysted stages. These cysts are 
oval and measure .01 by .007 millimeter. They are surrounded 
by a fairly thick hyaline layer through which the outline of the 
creatures can sometimes be seen quite distinctly. 

2. Teipaxosoma (see Plate I, e). — The tripanosomes, as 
usually seen in the blood of man, are elongated, unicellular, 
microscopic bodies, provided with a kind of fin-fold. The un- 
dulating membrane which runs along the dorsal edge of the 
body forms frill-like folds and terminates in a free whip-like 
filament (flagellum), which may vary greatly in length. 

Stained specimens show a large nucleus, usually situated at 
about the middle of the body, and a much smaller chromatin 
mass placed more or less near one pole. This chromatin mass 
is called the centrosome. The extremity of the body which 
encloses this is its anterior end. This organism varies greatly 
in length and shape in different stages of its development ; it 
may be pointed or obtuse. The posterior extremity always tapers 
and is continued into the free flagellum. Sometimes the body 
of the stained specimen show^s many darkly stained granules 
called chromatophores. (See page 100.) 

CLASS HI. SPOROZOA. 
Coccidia. 

1. Coccidioi Hovcixis or Perforaxs. — This organism is 
oval. The fertilized sporont stage is the oocyte (see Plate I, f), 
and measures .024 to .035 by .002 to .014 millimeter. It is sur- 
rounded by an integument with a double outline which has an 



PLATYHELMINTHES. 107 

opening at the pointed pole. The plasm, which is somewhat 
coarsely granular, entirely fills the integument or is gathered 
together in a rounded central mass. The coccidia are evacuated 
from the bowel in this stage, and spornlate in the open within 
two or three weeks. The fully developed sjwres are of a broad 
fusiform shape and measure .012 to .015 millimeter in length by 
.00? millimeter broad. They each contain two spovozoites, broad 
at one end and pointed at the other, forming a bent dumb-bell 
shaped body. The chromatin mass is called the centrosome, and 
the extremity of the body which encloses this, is the anterior 
extremity. 

A granular residual body lies in the concavity. This or- 
ganism is found in the intestinal tract, where it may give rise 
to violent auto-infection and chronic diarrhea. 

2. Hemospoidi^e. — These organisms are the cause of ma- 
laria and have been described in another section (see page 96). 

CLASS IV. INFUSORIA. 

Balaxtidium Coli or Paramecium Coli. — The body is 
oval, .06 to 1.0 millimeter in length, by .05 to .07 millimeter 
in width. The peristome is funnel-shaped or contracted, the 
anterior end being either blunt or pointed. The ecto- and ento- 
sarc are distinctly separate, the latter granular, and containing 
drops of fat, mucus, starch granules, bacteria, and occasionally 
leukocytes and erythrocytes. There are usually two contractile 
vacuoles. The anus opens at the posterior extremity, and the 
organism contains a macro- and a micro-nucleus. It occurs in 
the large intestines of man. 

B. PLATYHELMINTHES (Flat Worms). 

CLASS I. TREMATODES. 

1. Distomum Hepaticum (Liver fluke). — Length, 20 to 
30 millimeters; breadth 8 to 13 millimeters. The head-cone 
is 4 to 5 millimeters long, and sharply demarcated from the 
posterior part of the body. Spines in alternating transverse 
rows, and extending on the ventral surface to the posterior bor- 
der of the testes, and on the dorsal surface not quite so far. The 
spines are smaller on the head-cone than on the posterior part 



108 ANIMAL PARASITES. 

of the body, where they are discernible by the naked eye. The 
suckers are hemispherical and near each other. The oral sucker 
is about 1 millimeter, and the ventral about 1.6 millimeter in 
diameter. The pharynx; which includes almost the entire esoph- 
agus, measures .7 millimeter in length and .4 millimeter in 
breadth. The intestine bifurcates in the head-cone, and the 
branches are furnished with blunt sacs directed outwardly. The 
ovary is ramified and is situated in front of the transverse 
vitello duct. The shell-glands lie near the ovary and are in the 
median line. 

The ova are yellowish-brown, oval, with cap-like lid. They 
measure .130 to .145 by .07 to .09 millimeter. The average 
size is .132 by .08. The liver fluke is an inhabitant of the bile- 
ducts of man. 

2. Distomoi Pulmonale or Distomum Westermaxxi. — 
The body is of a faint, reddish-brown color and plump oval 
shape, with the ventral surface a little flattened. This organism 
measures 8 to 10 millimeters in length by 4 to 6 millimeters in 
thickness. It possesses two suckers of equal size (.75 millime- 
ter). 

The eggs are oval brownish-yellow, with a fairly thin shell 
and measure .0875 to .1025 millimeter in length, by .052 to 
.075 millimeter in breadth, the average being .0935 by .057 milli- 
meter. Their location is usually in the lung, but they may 
enter the blood-vessels and be carried to another part of the 
body. 

3. Distoviuvi Laxceolatuvi or Dicrocelium Laxceo- 
latum. — In the fresh condition this is a yellowish-red organism, 
the body is flat, almost translucent, with a conical neck at the 
level of the ventral sucker. This point is marked by a shallow 
constriction. The length and breadth vary according to the 
amount of contraction, being usually 8 to 11 millimeters by 
1.5 to 2.0 millimeters. The suckers are about one-fifth of the 
body-length distant from each other, and of about equal size 
(.23 by .25 millimeter). 

The eggs are oval with a sharply defined operculum at the 
pointed pole. They measure .030 by .011 millimeter, and occur 
in the feces. The parasites reside in the liver. 

4. Distomfm Hematobifvi or Bilharzia. — The male is 



PIATYHELMINTHE3. 101) 

whitish and measures L2 to 1 1 millimeters in length, hut is 

already mature when I millimeters long. The anterior end is 
(i millimeters or a little more in length; the suckers are situated 
near each other; the oral sucker is inf undibulif orm ; the dorsal 
lip being a little longer than the ventral. The ventral sucker 
is slightly the larger, and is pedunculated. A little behind the 
ventral sucker the body broadens to a width of 1 millimeter, 
decreasing, however, in thickness. The lateral edges curl ven- 
tral ly, so that the posterior part of the body is almost cylin- 
drical. The posterior end is somewhat attenuated, and the 
dorsal part of the posterior extremity of the body is covered 
with spinous papilli. 

The females are filiform, about 30 millimeters in length, 
pointed at each end and measuring .25 millimeter in diameter. 
The color of the body varies according to the contents of the 
intestine (posteriorly they are dark-brown or black). The 
cuticle is smooth except in the suckers, where there are very 
delicate spines, and at the tail end where there are larger spines. 
The anterior part of the body measures from .2 to .3 millimeter 
in length. 

The eggs are fusiform and much dilated in the middle. 
They have no lid and are provided with a terminal spine at the 
posterior extremity. The eggs measure 0.12 to 1.12 millimeters 
in length, by .05 to .073 millimeter in breadth. They are yel- 
lowish in color, slightly transparent, and provided with a thin 
shell. The spine may sometimes be absent. The eggs apparently 
vary greatly in size. The organism lives in the portal vein and 
its branches. 

CLASS II. 
a. Bothriocephaloidia. 

1. Bothriocephalic Latus or Tenia Lata. — Length 
from 2 to 9 or more millimeters. Color, yellowish gray; after 
lying in water lateral areas become brownish and the rosette 
of the uterus brown. The head is almond-shaped, 2 to 3 milli- 
meters in length. Its dorso-ventral axis is longer than its trans- 
verse diameter; the head is therefore generally flat, concealing 
the suctorial grooves at the borders. The neck varies in length 
according to the degree of contraction, and is very thin. There 



HO ANIMAL PARASITES. 

are from 3000 to -1500 proglottides. Their breadth is usually 
greater than their length, but in the posterior third of the 
body they are almost square, while among the very oldest some 
may be longer than they are broad. 

The eggs are large with brownish shells and small lids. 
They measure .068 to .071 by .0-15 millimeter. The proglottides 
near the posterior extremity of the worm are frequently eggless. 



b. Teniidae. 

1. Tenia Xana. — The worm is 10 to 15 millimeters in 
length, and .5 to .7 millimeter in breadth. The head is globular 
and is from .25 to .30 millimeter in diameters. The rostellum 
has a simple crown consisting of 24 to 30 hooks, which are only 
.014 to .018 millimeter in length. The neck is moderately long. 
The proglottides are very narrow, about 150 in number, .4 to .9 
millimeter in breadth, by .014 to .030 millimeter in length. 

The eggs are globular or oval, and measure .030 to .048 
millimeter. The oncospheres measure .016 to .019 millimeter 
in diameter. 

The worm lives in the intestines; the ova, and proglottides 
are found in the feces. 

2. Tenia Laxceolata or Hymenolepis Lanceolata. — 
The parasite measures 30 to 130 millimeters in length, and 5 to 
18 millimeters in breadth. The head is globular and very small, 
the rostellum is cylindrical with a crown composed of eight 
hooks (0.031 to 0.035 millimeter in length). The neck is very 
short. The segments increase gradually in breadth, but vary 
little in length. 

The ova have three envelopes and are oval, measuring 0.050 
by 0.035 millimeter. The external envelope is membranous and 
much wrinkled, the middle one is thick, and the internal one 
very thin. 

3. Tenia Solium or Tenia Vulgaris. — The average 
length of the entire tapeworm is about 2 to 3 meters, but 
may be more. The head is globular, 0.6 to 1.0 millimeter in 
diameter. The rostellum is provided with a double row of hooks, 
twenty-two to thirty-two in number; large and small hooks 
alternate regularly. The length of the largest hooks is 0.16 to 



PLATYHELMIXTHES. 1 1 1 

0.18 millimeter, of the small ones 0.11 to 0.11 millimeter. The 
average number of proglottides is 800 to 900; they increase 
very gradually in size. At about 1 millimeter behind the head 
they are square and have the generative organs fully developed. 
Segments sufficiently mature for detachment measure 10 to 12 
millimeters in length, by 5 to 6 millimeters in breadth. The 
fully devloped uterus consists of a median trunk with seven to 
ten lateral branches on each side, some of which are again 
ramified. 

The eggs are oval, the egg-shell very thin and delicate. The 
embryonal shell is very thick with radial stripes; it is of a pale- 
yellow color, globular, and measures 0.031 to 0.036 millimeter 
in diameter. The oncospheres with six hooks are likewise globu- 
lar, and measure 0.02 millimeter in diameter. 

4. Cysticercus Acanthotrias. — This resembles the cysti- 
cercus cellulose in form and size, but carries on the rostellum a 
triple crown each consisting of fourteen to sixteen hooks which 
differ from the hooks of the cysticercus cellulose or of the tenia 
solium by the greater length of the posterior root process and 
the more slender form of the hooks. The large hooks measure 
0.153 to 0.196 millimeter, the medium-size hooks 0.114 to 0.14, 
and the small ones 0.063 to 0.07. 

5. Texia Saginata or Tenia Mediocanellata. — The 
length of the entire worm averages 4 to 8 to 10 meters and 
more, even up to 36 meters. The head is cuboid in shape, 1.5 
to 2 millimeters in diameter. The suckers are hemispherical 
(0.8 millimeter), and are frequently pigmented. There is a 
sucker-like organ in place of the rostellum, and this is also 
frequently pigmented. The neck is moderately long and about 
half the breadth of the head. The proglottides average more 
than 1000, and gradually increase in size from the head back- 
ward. The detached mature segments are exactly like pumpkin- 
seeds — they are about 16 to 20 millimeters long, by 4 to 7 milli- 
meters broad. There are twenty to thirty-five lateral branches at 
each side of the uterus, and these often again ramify. 

The eggs are more or less globular, the egg-shell frequently 
remains intact, and carries one or two filaments. The embryonal 
shell is thick, radially striated, transparent and oval. It is 0.3 to 
0.4 millimeter in length, by 0.02 to 0.03 millimeter in breadth. 



112 ANIMAL PARASITES. 

This worm in the adult condition dwells exclusively in the 
intestinal canal of man. The corresponding cysticercus occurs 
in the ox and steer. 

6. Tenia Echixococcus. — This worm measures 2.5 to 5 or 
(J millimeters in length; the head is 0.3 millimeter in breadth, 
and has a double row of twenty-eight to fifty booklets on the 
rostellum. The size and form of these hooklets vary. The 
larger ones are 0.040 to 0.045 millimeter in length, the smaller 
ones are 0.030 to 0.038 millimeter. The suckers measure 0.13 
millimeter in diameter. The neck is short, behind which there 
.are only three or four segments, the posterior of which is about 
2 millimeters in length and 0.6 millimeter in breadth. The 
ovary is horse-shoe shaped, with the concavity directed back- 
ward. The median trunk of the uterus is dilated when filled with 
eggs, and instead of lateral branches has lateral protuberances. 
It is not uncommon for the eggs to form local heaps. The em- 
bryonal shell is moderately thin with radiating fibers, is almost 
globular, and measures 0.030 to 0.03 G millimeter in diameter. 

The mature parasite lives in the small intestine of the 
domestic dog and the wolf, and from them, the dog chiefly, they 
are transmitted to man. 

Diagnosis of Cestodes. — The microscopic examination of 
the feces should never be neglected when the presence of tape- 
worm is suspected. Often by careful, frequently repeated exami- 
nations, insistent symptoms, referable to the digestive tract, the 
nervous system or the general nutrition, may be cleared up by 
the finding of segments or ova in the feces. 

As the uterus of the tenia has no exit, the eggs can only 
find egress when the mature proglottide is injured. In the case 
of the tenia saginata the discharge of eggs is almost the rule. 
The proglottides when discharged are usually without eggs. The 
eggs of the solium and the saginata are only distinguished by 
their size. 

In examining the stools for evidences of tapeworms, one 
must be careful not to confound remnants of undigested food, 
mucous casts, and shreds of tendon with the proglottides. The 
proglottides, after being soaked in water, assume their character- 
istic form. As a rule the microscopic determination of ova is 
a more certain means of diagnosis than the macroscopic seg- 
ments. 



NEMATODES. 113 

To determine from the shape of the proglottide which 

variety of worm is present, it is advisable to fix the segment 
between two glass slides. The proglottide of the tenia solium is 
more delicate and more transparent than the tougher segments 
of the tenia Baginata. In the former the branching uterus is 

more plump, and the number of lateral twigs are from seven to 
ten, while the uterus of the tenia saginata shows from twenty to 
thirty or more. 

C. NEMATODES (Thread Worms). 

Stroxgyloides Intestinalis or Anguillula Intesti- 
NALIS et Stercoralis. — 1. (a) The parasitical generation 
(anguillula intestinalis) measures 2.3 millimeters in length by 
0.034 millimeter in breadth. The cuticle is finely transversely 
striated. The mouth is surrounded by four lips, the esophagus 
is almost cylindrical and is a quarter the length of the body. 
The anus opens just in front of the pointed posterior extremity. 

The eggs measure 0.050 to 0.58 millimeter in length, and 
0.030 to 0.03-1 in breadth. 

(b) The free-living generation (anguillula stercoralis) is sex- 
ually differentiated. The body of the male is cylindrical, 
smooth, somewhat more slender at the anterior extremity, and 
pointed at the tail end. The mouth has four lips and the 
esophagus a double dilatation. The males measure 0.7 by 0.035 
millimeter, and carry the posterior extremity curled up. The 
two spicules are small and much curved. The females measure 
1.0 millimeter in length or a little more, 0.05 millimeter in 
breadth. The tail end is straight and pointed. The yellowish 
thin-shelled ova measure 0.07 millimeter in length by 0.045 
millimeter in breadth. 

2. Filaria Sanguinis Hominis (filaria bancrofti or filaria 
nocturna). — The male is colorless and measures 40 millimeters 
in length and 0.1 millimeter in diameter. The cephalic ex- 
tremity is a little thickened, the posterior extremity is bent and 
rounded, but is not twisted cork-screw like. The anal orifice 
opens 0.138 millimeter in front of the posterior border. The 
female is brownish, 7.8 to 8.0 millimeters in length and 0.21 to 
0.28 millimeter in breadth. The cephalic and caudal extremi- 

8 



114 ANIMAL PARASITES. 

ties are rounded. Almost the entire body is occupied by the 
two uteri, from which the larvae emerge early. The length of 
the larvce average 0.13 to 0.3 millimeter, their breadth 0.007 to 
0.07 millimeter. They are surrounded by a delicate protective 
investing membrane which is not quite close to them. 

The lymphatic vessels in various parts of the body are 
doubtless the normal habitat of the adult worms, but these 
have also been found in the left ventrice of the heart. The 
young ova, by means of the lymph-stream, reach the blood and 
are distributed with it through the body. They also pass through 
the vessel walls and may be found in the fluid of the glands of 
the body. The larvae are first found in infected patients only in 
specimens of blood that have been taken after sunset. Their 
number increases considerably until after midnight, and after 
that time begin to diminish. From mid-day until evening no 
filariae are found in the peripheral blood. 

3. Trichocephalus Dispar or Ascaris Trichiura. — The 
male measures 40 to 45 millimeters in length, the spiculum is 
2.5 millimeters long and lies within a retractile pouch beset with 
spines. The female measures 45 to 50 millimeters in length, of 
which two-fifths appertain to the posterior part of the body. 
The ova are barrel-shaped and have a thick, brown shell which 
is perforated at the poles. Each opening is closed with a light- 
colored plug. The eggs measure 0.05 to 0.054 millimeter in 
length and 0.023 millimeter in breadth. They are deposited 
before segmentation. This worm usually lives in the cecum of 
human beings, and is occasionally found in the vermiform ap- 
pendix, in the colon, and exceptionally in the small intestine. 
Usually only a few are present, and they do not cause any par- 
ticular disturbance. 

The development of the eggs is completed in water or in 
moist soil, and occupies a longer or shorter period according to 
the season. The eggs and larvae possess great powers of resist- 
ance, and have been known to remain as long as five years in 
the egg-shell without losing their vitality. 

4. Trichixa Spiralis. — The male measures 1.4 to 1.6 mil- 
limeters in length and 0.04 millimeter in diameter. The ante- 
rior part of the body is narrowed, the orifice of the cloaca is 
terminal and lies between the two caudal appendages; behind 



NEMATODES. 115 

there are four papillae. The female measures 3 to 4 millimeters 
in length and 0.06 millimeter in diameter; the anus is terminal. 
Trichina spiralis occupies in its adult stage the small intestines 
of men and of various mammals, including the domestic rat, 
domestic pig and domestic dog. 

History of Development of Trichina Spiralis. — 
Shortly after entering the intestines the encysted trichinae 
escape from their capsules, and then enter the duodenum and 
jejenum where they become adult. During this period they 
do not greatly increase in size. The males grow from 0.8 to 1.0 
millimeter, the females from 1.5 to 1.8 millimeters. Soon after 
copulation, which takes place in the course of two days, the 
males die off, and the females, which soon attain the length of 
3.0 to 3.5 millimeters, either bore more or less deeply into 
the villi or penetrate the mucous membrane and enter the lym- 
phatic spaces. Here they deposit their young which, according 
to Leuckart, average at least 1500. The migrations are mostly 
passive, the larvae being carried along by the lymph-stream or 
by the circulating blood. The young brood is distributed 
throughout the entire body, but the conditions necessary to its 
further development, are found only in the transversely striated 
muscle. On the ninth or tenth day after infection the first 
trichinae have reached their destination, but further invasions 
are constantly taking place. Two or three weeks after infec- 
tion the spirally rolled up trichinae have grown to 0.8 to 1.0 
millimeter, and in their vicinity the muscle fibers are swollen. 
The capsule is formed by the inflamed connective tissue pro- 
ducing the cystic membrane. The cysts are lemon-shaped and 
usually lie with their longitudinal axis in the direction of the 
muscle fibers. On an average they measure 0A millimeter in 
length by 0.25 millimeter in breadth. 

5. Ankylostoma Duodenale or ITncinaria Duodenale. 
— The body is cylindrical, attenuated anteriorly, and of a 
slightly reddish color. In the oral cavity on the ventral surface, 
close behind the orifice, are four hook-like teeth directed back- 
ward; on the dorsal surface there are two teeth directed for- 
ward. The males measure 8 to 10 millimeters in length, and 0.-L 
to 0.5 millimeter in breadth. The bursa has two large lateral 
and one small dorsal alar processes. The females measure 12 to 



116 ANIMAL PARASITES. 

18 millimeters in length, and the caudal extremity has a small 
spine. The eggs are elliptical and have thin shells ; they meas- 
ure 0.032 to 0.0-45 millimeter in breadth, and 0.055 to 0.065 
millimeter in length. 

The ankvlostoma duodenale lives in the duodenum, and 
may rarely be found in the first part of the jejunum. 

6. Uxcixaria Americana can readily be distinguished 
from the preceding worm. It is shorter and more slender. The 
male worm measures from 7 to 9 millimeters in length by 0.3 
to 0.35 in diameter; the female 9 to 11 millimeters in length by 
0.4 to 0.45 millimeter in diameter. The buccal capsule is much 
smaller, and presents an irregular border; instead of four ven- 
tral hook-like teeth, it is provided with a vertical pair of promi- 
nent semilunar plates similar to those of a dog hook-worm. 
The pair of dorsal teeth is likewise represented by a pair of 
slightly developed chitinous plates of the same nature. 

The eggs are larger than in IT. Duodenale ; they measure 64 
to 75 micromillimeters by 36 by 40 micromillimeters in breadth. 
So far this worm has been found only in man ; its anatomical 
habitat is the small intestine. 

7. Ascaris Lumbricoides. — The coloring in the fresh 
stage is reddish- or grayish-yellow. The body is of an elongated 
spindle shape and the dorsal oral papillae carries two papillae 
of sense and the two ventral oral papillae are papillae of sense. 
The male measures from 15 to 25 centimeters in length and 
about 2 millimeters in breadth. The posterior extremity is 
conical and bent ventrically into a hook. The spicules measure 
2 millimeters in length and are curved and broadened at their 
free ends. On each side of the orifice of the cloaca are seventy 
to seventy-five papillae, of which seven pairs are post-anal. 

The female measures 20 to 40 centimeters in length 
and about 5 millimeters in diameter, the posterior extremity is 
conical and straight. The vulva is at the border between the 
middle and posterior thirds of the body, from which the two 
uterine tubes pass straight to the posterior end of the body. 
The convoluted ovaries measure ten times the length of the body. 

The ova are elliptical with a thick transparent shell and an 
external coating of albumin which forms protuberances. The 
ova measure 0.05 to 0.07 millimeters in length and 0.04 to 0.05 



NEMATODES. 117 

millimeters in breadth : they are deposited before segmentation. 
The albuminous coating is stained yellow by the coloring matter 
of the feces. 

This worm is one of the most frequent parasites of man, 
and is distributed over all parts of the world. 

s. Oxyuris Vermicularis ok Ascaris Vermicularis. — 

Color, white;" the attenuated cuticle forms swellings at the 
anterior end which extend some distance back along the middle 
o\' the ventral and dorsal surfaces. There are three small re- 
tractile labial papilla? around the mouth. The male measures 
3 to 5 millimeters in length, and shortens on death. The poste- 
rior extremity of the body is rolled ventrally and presents pa- 
pilla?. The female is 10 millimeters in length and 0.6 milli- 
meter in diameter. The anus is about 2 millimeters in front 
of the tip of the tail ; the vulva is in the posterior third of the 
hody. The eggs are oval, thin-shelled, and measure 0.05 by 
0.02 millimeter. They are deposited with embryos already de- 
veloped, and are seldom found in the feces. 



IX. 

DETERMINATION OF THE FUNCTIONS 
OF THE STOMACH. 



THE GASTRIC CONTENTS. 

The significance of the term gastric contents, in the fol- 
lowing pages, is taken to mean the material found in the stomach 
and extracted by the gastric tube at the expiration of a fixed 
and definite period after the ingestion of a test-breakfast. 1 

THE VOMITUS. 

It is not good practice to utilize vomitus for the purpose 
of chemical analysis. Such material is usually of very uncertain 
composition, being contaminated with mucus from the upper 
part of the tract; further, the amount or composition of the 
food previously ingested is an unknown factor, affecting mate- 
rially the quantitative and qualitative findings. However, it 
may be found of decided advantage to test all vomited matter 
for the presence or absence of acidity and free hydrochloric acid. 
In the presence of suspected cancer the vomited material may 
be searched for necrotic tissue shreds or sarcinae. 

METHODS OF OBTAINING SPECIMEN FOR 
EXAMINATION. 

The usual apparatus employed to remove the test-meal 
comprises the well-known gastric tube of soft red rubber, fitted 
at one end with a soft rubber funnel, and containing near this 
a bulbous expansion without valves. A recent modification of 
and improvement over this is the large bulb devised by Ewald. 
This bulb is sufficiently large to contain the total quantity of 



1 The Ewald test-breakfast consists of an ordinary roll weighing about 35 grams, 
and 300 centimeters of water or weak tea without milk or sugar. 



(118) 



METHODS OF OBTAINING SPECIMEN. 



119 



material removed, thus overcoming in a measure the difficulties 
of the smaller bull). These two methods are usually successful 
in obtaining the desired material for examination, but are dif- 
ficult to manage and possess a decided disadvantage in that they 




Fig. 15.— Complete Outfit for Gastric Test-meal Removal, 
Lavage an"d Inflation. 



do not provide for the important procedure of lavage and infla- 
tion. 

Also, tubes of irregular caliber are difficult to cleanse, 
and unless great care is exercised in this direction, they may 
become a source of contamination, if not carriers of infection. 



120 THE STOMACH. 

Some years ago Dr. Judson Daland adopted for this pur- 
pose two large open-mouth bottles and a double-action Davidson 
bulb; these are used in conjunction with the plain gastric tube. 
With this arrangement, aided by an assistant, it is possible to 
rapidly and cleanly obtain a sample of gastric contents (in con- 
siderably less than a minute), and to follow immediately with 
lavage and inflation if desired. 

To those who have used this method with uniform success 
in both private and hospital practice, its advantage is evident, 
the only drawback being the almost absolute necessity for trained 
assistance, this being occasioned by the complicated nature of 
the apparatus, which requires a second pair of hands to manipu- 
late the bottle and its connections during the passage of the tube. 
To overcome this drawback the author has devised a reversing 
valve (see Fig. 15). With this valve it is unnecessary to make 
any change in the tube connections when once the apparatus is 
set up. The perfected device has been worked out with great 
care, having in mind the necessity of a simple mechanism capa- 
ble of being cleansed and kept in repair without difficulty. 



PRELIMINARY PREPARATION OF THE PATIENT. 

The preliminaries leading up to the extraction of the test- 
meal should be as nearly uniform in every case as possible. By 
adopting a definite routine and adhering to it we eliminate, in 
a great measure, the errors which would otherwise creep in and 
lesson the value of the findings. The adoption of the following 
rules will enable the examiner to obtain a series of reports in one 
case or in groups of cases of far greater value than could be 
obtained by an irregular technic : — 

First. — It is advisable that all medication should be with- 
held until after the gastric analysis has been made. If this is 
impossible, then a period of days should elapse before making 
the test, during which all treatment is stopped. 

Second. — The test-meal should be given on an empty stom- 
ach; either one which has been emptied by fasting or by pre- 
vious lavage. 



MODIFIED EWALD BREAKFAST. 121 

Third. — The volume and composition of the test-meal 
should he uniform. The particular meal employed to be deter- 
mined by the operator according to his preference. 

Fourth. — The removal of the meal should (depending on 
its composition) be accurately timed; this time to be measured 
from the beginning and not from the end of the meal. 

Fifth. — The test-meal should be removed without dilution 
if possible. If dilution should be necessary to accomplish re- 
moval, the amount of water used should be definitely known, 
and the total withdrawn must exceed the amount used. If this 
rule is not observed, all investigations of a quantitative nature 
are valueless. 

Sixth. — The tube should remain within the esophagus only 
long enough to allow time to compress the bulb. If much time 
elapses after introduction before removal of the contents, the 
hypersecretion of mucus occasioned by the presence of the tube 
will alter the composition of the sample and possibly result in 
erroneous conclusions. 

A tube of relatively large diameter is to be preferred, being 
easier of insertion, less uncomfortable to the patient, and in- 
suring greater success in obtaining a sample. A suitable tube 
should measure between three-eighths and one-half inch outside 
diameter, and should be provided with both a terminal and lat- 
eral opening in the gastric end. 

A lubricant for the tip of the tube is unnecessary. The 
contact of the tube with the patient's pharynx immediately ex- 
citing sufficient flow of mucus for this purpose. This is prefer- 
able to either nauseating oil or hygroscopic glycerine. In case 
of great reflex excitability of the pharyngeal constructors, this 
region may be first sprayed with a dilute solution of cocaine. 

Modified Ewald Breakfast. — Of the test-meals employed, 
one of generally useful composition includes the white of two 
eggs poached or soft boiled, without yelks and without season- 
ing; two pieces of toast without butter (the slightest trace of 
butter or fat will cause lactic and butyric acid fermentation), 
and a cup of tea without milk or sugar. 

This meal should be removed at the expiration of one hour 
when, under ordinary circumstances, there will be recovered be- 
tween 30 and 90 cubic centimeters. 



122 THE STOMACH. 

The component parts of the apparatus are as follows: — 

A plain gastric tube without bulb or funnel. 

A double-action Davidson hand-bulb. 

A large rubber stopper having two perforations. 

Two wide-mouth bottles containing more than a liter each, 
and graduated in cubic centimeters. 2 

A short length of large glass-tubing and some one-quarter 
inch rubber-tubing. 

The reversing valve. 

Technic of Removal. — Fill one graduated bottle to the 500 
cubic centimeter mark with warm, sterile water. Fit the double 
perforated stopper with the glass tube and the reversing valve. 
Place the stopper firmly in the empty bottle and attach the gas- 
tric tube to the glass tube, and the two ends of the double action 
bulb to the two horizontal tubes of the valve. Finally, ascertain 
the direction of the air-current through the valve by making a 
few pressures on the bulb. Set the valve to make negative pres- 
sure within the bottle. 

The tube should now be passed, with the patient preferably 
in the sitting posture. As soon as the tube enters the cardiac 
orifice, a few quick pressures are made on the bulb. This de- 
velops a slight degree of negative pressure within the bottle, 
when the gastric contents will immediately flow into the bottle. 
Sudden stopping of the flow from occlusion of the tube by par- 
ticles of food or mucus, may immediately be removed by mo- 
mentarily reversing the lever. This will cause a small portion 
of the gastric contents to return through the tube, effectually 
washing out the obstruction. This simple maneuver may be 
repeated as frequently as necessary to obtain a sufficient speci- 
men. 

In the event of failure to obtain sufficient material by this 
means, the difficulty will usually be overcome by the introduc- 
tion of a measured amount of water. To accomplish this the 
stopper with all its connections is removed from the bottle and 
fitted into the bottle containing the 500 cubic centimeters of 
warm water. The valve is set to make positive pressure within 



2 These special bottles are not necessary for practical purposes ; any large, open 
mouth bottle of sufficient quantity, such as a quart milk bottle, may be substituted. 
A mark must be made at the measured 500 cubic centimeter mark on one bottle. 



INFLATION OF THE STOMACH. 123 

the bottle, and by means of the bulb about 400 cubic centimeters 
are run into the stomach; then by reversing the valve the whole 
is withdrawn. It is necessary to recover more than a total of 
500 cubic centimeters if any determinations of a quantitative 
nature are to be made. 

If it has been necessary to resort to the introduction of 
water to effect the removal of the test-meal, allowance must then 
be made in the final calculations, so that the results will repre- 
sent the quantities in pure gastric contents. 

For Example. — Suppose after employing 500 cubic centi- 
meters a total of 550 cubic centimeters is recovered, of this 
amount only 50 cubic centimeters represent actual gastric con- 
tents, or one part in every eleven. It will be necessary then to 
multiply any figures obtained in the calculations of acidity by 
the factor eleven in order to express the results in terms of 
undiluted gastric contents. 

Inflation of the Stomach. — Next to the x-ray, probably the 
best method of determining the size, shape, and location of the 
stomach is by the introduction of air. To accomplish this two 
methods are available. Of these, the second is greatly to be 
preferred for reasons to be stated later : — 

1. The first consists in administering one dram of sodium 
bicarbonate dissolved in a little water, to be immediately fol- 
lowed by an equal quantity of tartaric acid, also in solution. 
The combination of these causes the evolution of carbon dioxide 
gas within the stomach, which immediately distends that organ. 
This method is open to serious objection, because the quantity 
of gas produced cannot be controlled and over-production, be- 
sides causing great discomfort if not doing actual damage, may 
result in hemorrhage and great cardiac embarrassment. On 
the other hand, sufficient gas may not be evolved to completely 
distend the stomach, and thus its full size and shape fail to be 
accurately determined. In the light of these facts it would 
seem best that this method be permanently abandoned for the 
following, which is more in accord with the principles of scien- 
tific medicine. 

2. The second method of inflation is accomplished through 
the stomach tube by means of a Davidson bulb. This simple 
combination may be employed with safety and accuracy, and 



124 THE STOMACH. 

even in the absence of the graduated bottles and reversing valve 
can be made to serve. 

A greater refinement in the technic is attainable with the 
aid of bottles and valve above described. Their use may con- 
veniently follow the removal of the test-meal. With this outfit 
in addition to inflation we may also roughly measure the cubic 
contents of the distended stomach, by introducing a measured 
quantity of air. 

Technic of Inflation. — By inflation the position of the stom- 
ach may be accurately outlined. After the test-meal has been 
removed the patient is placed in the semi-recumbent posture, 
and the empty stomach is outlined as accurately as possible by 
auscultatory percussion. Air is then introduced into the stomach 
through the tube in sufficient quantity to produce a distinct 
change in the auscultatory percussion-note. The quantity of air 
required to accomplish this is quite small, not sufficient to alter 
the relation of the organ to surrounding viscera, as is the case 
when the stomach is ballooned with air. By the proper working 
of the valve and the bulb on the apparatus, we can change the 
gastric note at will, thereby being able to differentiate abso- 
lutely between gastric and colonic tympany. 

By this method gastroptosis can be absolutely determined, 
the author having in many instances had the results confirmed 
by the x-ray. 

Coxtraixdicatioxs to Ixflatiox. — These are the same as 
for the use of the stomach-tube itself, viz. : Myocardial degen- 
eration, with or without endocarditis ; angina pectoris, aneurism, 
advanced vascular degeneration, hemorrhage from any part of 
the body, and all diseases in which hemorrhage is likely to occur. 
Gastric ulcer is, of course, a contraindication, especially when 
hemorrhage has been noted. Another contraindication, and one 
which is not sufficiently emphasized, is neurasthenia and allied 
mental states with gastric symptoms. In these diseases the 
symptoms referable to the stomach are not due to organic change 
in that organ, but are psychic in origin. The use of the tube 
in these cases serves but to reinforce the idea of disease of the 
stomach, and renders a cure more difficult and occasionally im- 
possible. This, of course, refers to those cases which are ob- 
viously psychic in origin. In certain cases it is not possible to 



INFLATION OF THE STOMACH. 



125 



determine whether the disease is organic or not; here the tube 
would have to be used for diagnostic purposes. It has been sug- 
gested that the unpleasantness of the passage of the tube itself 
will act as a curative agent. Experience teaches that usually 
more harm than good will come from the use of the tube with 
this intent. 

To Determine the Capacity of the Stomach by Inflation. — 
To accomplish this the apparatus employed in the preceding de- 




_S-0 



c.c 



JZl 



c.c. 



Fig. 16.— Diagrammatic Representation of Arrangement of Bottle 
for Measuring Cubic Contents of Stomach. 



seription must be augmented by the addition of a second double- 
perforated rubber stopper and a second section of glass-tubing, 
arranged in a similar manner to the first. 

The apparatus is*to be set up as follows (see Fig. 16) : The 
extremity of the gastric tube is attached to the short glass-tube 
<>f the second stopper, and a section of tubing made to join the 
long glass tube of each stopper. The bottle belonging to the 
valve and bulb is filled with water to the 1000 cubic centimeter 
mark, and both stoppers placed in their respective bottles. Now, 
with the tube in the stomach and the valve set for compression. 



126 THE STOMACH. 

the water is gradually forced over into the second bottle. This 
in turn displaces the air, and forces it into the stomach. When 
the patient indicates that the stomach is full, the amount of 
water displaced from the first bottle will equal the amount of 
air forced into the stomach, and can be read in cubic centimeters 
from the scale on the bottle. (See diagram.) The valve should 
now be reversed and the air withdrawn. 

Technic of Gastric Lavage. — Prepare a large pitcher of 
sterile water at body temperature, and a receptacle for waste. 
If desired normal saline or dilute alkaline water may be sub- 
stituted. Place 500 cubic centimeters of the wash-solution in one 
bottle, and by means of the valve and bulb force a few hundred 
centimeters into the stomach ; then, after allowing it to remain 
for a few moments, reverse the valve and withdraw as much as 
possible. Discard the returned water and repeat this process 
until the wash-water returns clear. Finally, a quantity of this 
saline or alkaline solution may be allowed to remain when the 
tube is withdrawn. 

Indication for the Use of the Gastric Tube. — The tube is 
positively indicated in cases of carcinoma associated with py- 
loric stenosis. The result here is decided relief from distressing 
symptoms, and often actual prolongation of life. 

In certain cases of pernicious anemia, especially in the later 
stages, stagnation of food gives rise to distressing symptoms, and 
by allowing absorption of toxins, hastens the progress of the 
disease. 

Finally, in cases where poison has been taken into the stom- 
ach in toxic closes, the stomach tube is often of great service in 
affording prompt removal. 



THE ACIDS OF DIGESTION. 

Under conditions of health, after from ten to fifteen min- 
utes following the ingestion of food the gastric contents are 
acid, due to the presence of free acids or acid salts. At this 
time the free acid recognized is lactic acid. Up to thirty-five 
or forty minutes lactic acid predominates, and only traces of 
HC1 can be detected. Shortly after this the lactic acid disap- 



ACIDS OF DIGESTION. 127 

pears and only HC1 remains, so that at the end of one hour no 
Lactic aeid can be demonstrated. 

Hydrochloric acid is actually present from the beginning, 
but its presence is masked by the excess of lactic acid and the 
HC1 combined with bases. Free HC1 increases with the progress 
of digestion until it reaches 0.15 to 0.20 per cent, after a light 
meal, or from 0.20 to 0.33 per cent, after an abundant meal. 



CHEMICAL COMPOSITION OF THE GASTRIC JUICE 
(containing water with saliva). 

Water 994.40 parts. 

Solids 5.60 

Organic material 3.10 

Mineral salts 2.50 

Sodium chlorid 1.46 

Calcium chlorid 0.16 

Potassium chlorid 0.55 

Ammonium chlorid Trace. 

Calcium phosphate Trace. 

Magnesium phosphate Trace. 

Iron 0.12 

Free HC1 0.20 



FREE ACIDS. 

For the simple qualitative demonstration of free acids, 
organic and inorganic, the congo-red and tropeolin papers are 
convenient. The former turns dark-blue, the latter dark-brown, 
when moistened with a solution containing free acids, but neither 
of these react to acids which are combined with bases. 

Detection of Free Hydrochloric Acid. — For the qualitative 
determination of the presence of free HC1, a number of tests 
are available. 

Tests. — 1. Topfer's Dimethyl-amido-azobenzol : For 
this test either the 0.5-per-cent. alcoholic solution of this chem- 
ical can be used, or for convenience filter-paper may be soaked 
in the 0.5-per-cent. solution, allowed to dry, and then kept bot- 
tled for use. This solution is delicate for 0.003 per cent. HCL 
Combined HC1 as well as acid salts and inorganic acids, in the 
concentration in which they occur in the stomach, will not 



128 THE STOMACH. 

cause this solution or the prepared paper to become pink. A 
pink reaction denotes the presence of free hydrochloric acid. 

2. Guxzberc/s Phloroglucin Vanillin (for reagent see 
Appendix). — This reagent, if active, is of a pale-yellow color. 
It darkens and deteriorates on exposure to light, so should be 
kept in a dark-colored bottle or prepared fresh each time that 
it is required. A drop or two of this reagent is placed in a 
porcelain dish, together with an equal amount of filtered gastric 
contents, and heat gently applied until the liquid has evapo- 
rated. If HC1 be present a rose-red color will appear at the 
margin of the evaporating fluid. This test is unmistakable and 
is most delicate, demonstrating the presence of free HC1 in the 
proportion of 1 to 10,000 or 0.005 per cent. The reaction is 
not interfered with by albuminates, by salts present in the 
usual amount, or by organic acids. 

QUANTITATIVE ESTIMATION OF TOTAL ACIDITY. 

The reaction of the filtered gastric juice being determined 
by the congo-red or the tropeolin papers, the total acidity is next 
determined by titrating against a decinormal sodium hydrate 
solution. 

Technic. — A Mohr's burette is filled to the "0" mark with 
standardized decinormal sodium hydrate solution. 

To 10 cubic centimeters of filtered gastric juice in a por- 
celain dish, 10 cubic centimeters of distilled water is added, and 
one or two drops of a 1-per-cent. alcoholic solution of phenol- 
phthalein added as an indicator. (This indicator is inactive in 
the presence of carbon dioxid.) In the presence of free acids 
this mixture is colorless. The sodium hydrate is now run in 
slowly with constant stirring, until the rose-color, which appears 
on the addition of each drop of alkaline solution, no longer dis- 
appears nor is intensified by further addition of the sodium 
hydroxid solution. 

As a rule the acidity of the gastric contents, one hour after 
the ingestion of the test-breakfast, requires from -1 to 6 cubic 
centimeters of decinormal XaOH to neutralize 10 cubic centi- 
meters of gastric contents. For example, suppose 5.2 cubic 
centimeters were required to neutralize 10 cubic centimeters of 



FREE HYDROCHLORIC ACID. 129 

gastric contents (this is within normal limits). The result of 
the estimation is usually expressed in parts of decinormal NaOH 
per 100 parts of gastric filtrate. This expression is easily ob- 
tained by moving the decimal point one place to the right. The 
result in the above example would therefore be recorded as 52 
parts of decinormal NaOH per 100 parts of gastric contents. 

QUANTITATIVE ESTIMATION OF FREE HYDRO- 
CHLORIC ACID. 

To 10 cubic centimeters of filtered gastric juice add an equal 
quantity of distilled water, and one or two drops of Topfer's 
reagent. This mixture placed in a porcelain dish is titrated with 
a decinormal solution of NaOH until the pink color of the solu- 
tion has been entirely removed and is replaced by a pale-yellow. 
Suppose, for example, 6 cubic centimeters was the amount 
required to neutralize the free HC1 contained in 10 cubic centi- 
meters of gastric contents. To obtain the percentage of HC1 in 
the gastric contents the following calculations are required : — 

One centimeter of decinormal NaOH solution is equivalent 
to 0.00365 gram of HC1 (decinormal NaOH is equivalent to 4 
grams of NaOH dissolved in exactly 1000 cubic centimeters of 
distilled water; each 1 cubic centimeter of this solution should 
exactly neutralize 0.00365 gram HC1). Therefore 6 x 10 would 
equal the number of cubic centimeters of NaOH solution required 
for every 100 cubic centimeters of gastric contents, and the 
result, 60 x 0.00365, would equal the percentage of HC1 in the 
specimen under examination, w r hich would be 0.219 per cent. 
HC1. 

Causes of Lowered Gastric Secretion. — I. In acute and 
chronic gastric inflammation. 

II. Atrophy of the gastric mucosa from any cause. 

III. General functional depression. 

IV. Expression of a gastric neurosis. 

V. Congenital peculiarity. 

DETERMINATION OF ORGANIC ACIDS. 

These include lactic and acetic, and the true fatty acids, 
particularly butyric. Acetic and fatty acids are not found dur- 



130 THE STOMACH. 

ing normal digestion, and if present, as they sometimes are, have 
either been introduced with the food or have been produced by 
fermentation of carbohydrates set up in the stomach by bacteria 
introduced with the saliva. 

The physiologic presence of lactic acid during the first stage 
of gastric digestion, may be due either to its formation within 
the stomach, or from its having been introduced with the food, 
as in baked bread. 

Ufflemax's Test for Lactic Acid. — The addition of a 
few drops of filtered gastric contents to Uffleman's reagent (see 
Appendix) in a test-tube will, in the presence of lactic acid, 
change the amethyst-blue to a canary-yellow. This test is posi- 
tive in the presence of 1 part of lactic acid in .20,000. 

Sources of Error. — Lactates cause the same reaction. This 
is, however, immaterial, since we desire to recognize lactic acid 
whether free or combined. The reaction also takes place with 
alcohol, sugar, and certain salts, particularly the phosphates, but 
rarely in their usual concentration after the test-breakfast. 

The fatty acids, particularly butyric, strike a tawny-yellow 
color with a reddish tinge with Uffleman's reagent. The reac- 
tion is positive with 1 part in 2000. Fatty acids may also be 
detected by heating to boiling a few cubic centimeters of gastric 
filtrate in a test-tube, over the mouth of which a strip of moist 
neutral litmus paper is placed. On this the vaporized volatile 
acid will produce the usual change. 

Acetic acid is easily recognized by its odor, but it may 
also be detected by neutralizing a water}' residue after the re- 
moval of an ethereal extract, with sodium carbonate, and then 
adding neutral ferric chloricl solution. In the presence of 
acetic acid a striking blood-red color will be produced. A simi- 
lar reaction occurs in the presence of formic acid, but this acid 
is never present in gastric juice. 

Alcohol. — This is sometimes formed in the stomach during 
intense yeast fermentation. This may be detected by the Lis- 
ten s iodoform test, applied as follows: To a portion of the 
gastric filtrate add a small portion of liquor potassii, and then 
a few drops of a solution of iodine and potassium iodide in water. 
(See Appendix.) If alcohol be present a yellowish scum gradu- 
ally occurs on the surface, which is readily recognized as iodo- 



TESTS FOR OCCULT BLOOD. 131 

form by its odor. The same reaction occurs in the presence of 
acetone, but occurs more rapidly. 

Propeptone and Peptone. — These are products of albumin 
digestion, and when present indicate the activity of this part 
of the gastric function test. A few drops of filtrate added to 
hot Fehling's solution produces a purplish color, if they are 
present. 

Starch. — The addition of a drop of LugoPs solution to a 
piece of filter paper upon which some unfiltered gastric contents 
has been allowed to fall will, in the presence of starch, yield a 
blue reaction. This reaction is intensified by the addition of a 
drop of crude nitric acid. 



MICROSCOPIC EXAMINATION. 

Examination of the unfiltered gastric contents under low 
power will reveal the excentrically marked oval starch grains, 
the budding yeasty cells, food debris, and epithelial cells, # etc. 
Under pathological conditions we may find a great excess of 
epithelial cells, shreds of mucous membrane or fragments of 
new growths, various bacilli and sarcinse, pus-cells, blood-cells, 
and bacteria. 

TESTS FOR OCCULT BLOOD. 

Blood may be intermittently present in the gastric contents 
or gastric vomitus as a result of one of the following condi- 
tions : — 

I. Ulcer of the stomach or intestines. 

II. Benign pyloric stenosis. 

III. Spasm of the pylorus. 

Occult blood is usually permanently present in cases of 
malignant disease of the stomach or esophagus. 

Boas Modification of the Weber Test. — To 15 cubic 
centimeters of gastric contents, add an equal amount of ether 
and thoroughly agitate. To this mixture add 3 to 5 cubic centi- 
meters of strong glacial acetic acid, and again agitate. ^N"ow 
allow the mixture to settle and decant 10 or 15 cubic centimeters 
of the clear supernatant liquid, and to this add 4 or 5 cubic 



132 THE STOMACH. 

centimeters of ozonized oil of turpentine. In the presence of 
blood a violet or blue color will appear, which is intensified by 
the addition of chloroform. (See also test on page 154.) 

Acetic Acid Ethek-Guaiac Test. — To 10 cubic centi- 
meters of gastric contents add 10 cubic centimeters of ether and 
5 cubic centimeters of strong acetic acid. Thoroughly shake, 
and add two or three grains of powdered gum guaiac, and again 
agitate ; allow to settle, and then add a few drops of fresh solu- 
tion of hydrogen dioxicl. In the presence of blood a purple or 
blue ring will appear at the line of contact, or the solution grad- 
ually assume a grayish-blue color. 

ESTIMATION OF PEPTIC ACTIVITY. 

The digestive power of the filtered gastric contents from the 
recovered test-meal depends upon, first, the amount of pepsin 
contained, and, second, upon the amount of free acid present, 
particularly the free hydrochloric acid. Artificial digestion is 
the only means at our command by which we may determine the 
peptic activity of the gastric juice. 

Method oe Ewald. — Prepare from the albumin of eggs, 
which have been boiled just sufficiently to cause firm coagula- 
tion, small discs or squares by first cutting thin slices of the 
coagulated albumin, and from these slices the cubes or discs. 
These bits of albumin may be prepared in bulk and preserved 
in glycerin, which is carefully washed off before using. 

The Test. — Place an equal quantity (5 to 8 cubic centi- 
meters) of filtered gastric juice in each of four test-tubes, add 
to each tube one or two pieces of the prepared albumin ; then, 

To tube 1 add nothing. 

To tube 2 add one drop of HC1. 

To tube 3 add four grains of pepsin. 

To tube 4 add the above quantities of HC1 and pepsin. 

These tubes are now placed in an incubator at 37° C, and 
from time to time examined to note the progress of liquefaction 
of the albumin. By comparison we can roughly determine 
whether digestion is progressing normally in the unaltered tube, 
and whether pepsin, hydrochloric acid, or both, are necessary to 
accomplish it. 



ESTIMATION OF PEPTIC ACTIVITY. 133 

The results obtained from this investigation can only be 
considered in the light of an approximate indication of the peptic 
activity of the material tested. 

According to Xierenstein and Schiff 3 it is necessary to dif- 
ferentiate three factors in order to arrive at an adequate idea 
of the significance of these pepsin determinations. These are: 
the diluting secretion, the pepsin secretion, and the hydrochloric 
acid secretion, all three of which must be regarded as distinct 
expressions of the secretory activity of the glandular parenchyma. 

Following a better knowledge of the activity of these dif- 
ferent factors, two improved modifications of the method of 
pepsin determination have been advanced. These are the meth- 
ods of Hammerschlag and of Mett. 4 

Method of Hammerschlag. — Briefly outlined the method 
depends upon the activity of a few centimeters of gastric juice 
upon a 1-per-cent. filtered solution of egg albumin. Two test- 
tubes, one filled with the albumin solution alone and the other 
with the albumin solution, plus the gastric filtrate, are incu- 
bated for an hour at 37° C. At the expiration of this time the 
albumin content of each tube is estimated volumetrically, ac- 
cording to the method of Esbach (see page 198). The differ- 
ence between the albumin precipitate in the two tubes is equal 
to the amount of albumin digested, and therefore is a measure 
of the peptic activity of the gastric juice. The objections to this 
method are, that the Esbach method is not very accurate, and 
also that albumoses are partly precipitated by the reagent. How- 
ever, this method is sufficiently accurate for the ordinary clinical 
purposes, and when the Mett method cannot be followed, may 
be adopted. It would be better, however, to employ a method 
which employs diluted gastric juice, as in the Mett method. 

Mett's Method of Peptic Determination. — Glass ca- 
pillary tubes from 1 to 2 millimeter 5 diameter and of convenient 
length, are filled by suction with the fluid portion of egg albu- 
min. In order to avoid accidental variation in the egg albumin, 
it is better to use the albumin of several eggs mixed. These 
tubes as they are filled should have their ends plugged with 



3 Arch. f. Verdauungskrankh, Vol. iii, 1902. 

4 Sahli's Diagnosis. 

5 J. A. D., Petersburg, 1889, from Palow's Laboratory. 



134 THE STOMACH. 

bread crumbs to prevent loss of contents before coagulation. 
After filling they are placed horizontally in a simmering water 
bath, where they are allowed to remain for from four to five 
minutes. While boiling, the tubes should be kept in motion to 
insure uniform coagulation of the albumin. When freshly made, 
the albumin tubes contain innumerable bubbles which, however, 
gradually disappear, after four or five days the tubes are ready 
for use. i-Vfter boiling, the tubes are wiped dry and the 
ends closed with sealing wax or stick lac to prevent drying. 
Thus prepared a stock of tubes may be kept indefinitely, but it 
should be ascertained before using that the albumin is still 
evenly in contact with the walls of the tube ; if they have dried 
out they are not fit for use. 

These tubes are cut as needed into lengths of from 2 to 3 
centimeters. This cutting is best accomplished by nicking the 
glass with a small triangular file, when a quick bend at this 
point will usually cause accurate fracture of both tube and albu- 
min. Portions containing air bubbles or showing irregular 
fracture, should be discarded. 

Teclinicfi — For quantitative determinations of the peptic 
activity of the filtered gastric contents, employ a dilution of 
1 to 16, using as a diluent ^ HC1, using for each test 1 cubic 
centimeter of contents and 15 cubic centimeters 20th normal 
HC1. 

This mixture is placed in a covered Stender 7 dish after two 
Mett's tubes have been placed in it, and the specimen allowed 
to digest in an incubator (37° C.) for twenty-four hours. At 
the end of this period the digested cylinders of egg albumin at 
each of the four ends are measured off and the average reading 
calculated. The relative amount of pepsin is then obtained by 
squaring the result (Shurtz's law), and if desired multiplying 
by the dilution, e.g., 16. For making the measurements a pair 
of callipers with" a vernier reading 0.1 of a millimeter, is con- 
venient. A lens is useful principally for reading the vernier. 
One end of the tube is placed against one jaw of the calliper, 
and the other is separated until its end is just visible through 



6 After Sailer and Fair, U. of Pa. Med. Bui., Oct. 1906. 

7 A Stender dish is similar to a Petri dish, but is deeper, with a flat grround 
grlass lid. 



ESTIMATION OF PEPTIC ACTIVITY. 135 

the opalescent edge of albumin. H the tube is at all oblique the 
shortest side is taken, while if the albumin is at all uneven, the 
highest point to which digestion has extended is taken. In place 
of the callipers the ordinary mechanical stage, which is usually 
fitted with a vernier scale, may be used. 

Xierenstein and Schiff have proven that a dilution of at 
least 1 to 16 is absolutely necessary if we wish to obtain a rela- 
tive idea of the quantity of pepsin as estimated from the diges- 
tion length. This is because w r ith a dilution of less than 16, 
the length of digestion decreases as the length of time increases, 
owing to the concentration and activity of the inhibiting sub- 
stances. This great dilution also decreases the amount of pepsin 
in the mixture, so that the digestion length is kept within the 
limits of Shurtz^s law r (3.6 millimeters in twenty-four hours). 
As there are instances w^hen this dilution is not sufficient (with 
very active pepsin solutions), it may become necessary when 
the digestion length exceeds 3.6 millimeters with the 16-fold 
dilution, to repeat the pepsin test w r ith a dilution of 1:32; then 
by squaring the length as before, and multiplying this quantity 
by 32, the relative amount of pepsin in the undiluted gastric 
contents is obtained. It is clear that in this estimation the unit 
of the relative amount of pepsin will be that quantity of pepsin 
by which 1 millimeter of albumin in the Mett tube will be 
digested in twenty-four hours with an acidity of 0.18 HC1. In 
this determination we do not consider the absolute quantity of 
pepsin, but simply the degree of concentration, for the result of 
Mett's method is the same whether large or small quantities of 
digestive substances are employed. 8 

Summary. — Conclusions of Sailer and Farr regarding varia- 
tions due to alterations in technic 9 : — 

1. Difference in the caliber of the tubes. The variations in 
the reading of the two ends of one tube were as great as between 
tubes of different caliber for tubes between 1 and 2 millimeters. 
In tubes greater than 2 millimeters the rate of digestion seemed 
to be uniformly greater in tubes of larger diameter. 

2. The age of the tubes. Provided there is no separation 



8 Sahli's Diagnosis, fourth edition, 1905. 
9 Sailer and Farr, he. cit. 



136 THE STOMACH. 

of the albumin nor putrefactive softening, age does not seem to 
affect the tubes. The method of preparation tends to make them 
sterile. Tubes .that are too fresh (less than four or five days 
old) are said to digest more rapidly than tubes that are "ripe." 

3. Variability in the digestion of albumin from different 
eggs is an unimportant factor. 

4. The degree of digestion within certain limits is said to 
vary directly with the duration of digestion. 

5. Variations in the temperature of the incubator. This is 
undoubtedly an important factor, but there are no definite data 
to offer. 

6. The effect of variations in the acidity of the original 
specimen. This has been provided against in the technic of the 
method, since the use of so large an amount of diluting acid 
solution tends to render the acidity of the diluted specimen 
uniform in every instance. 



ESTIMATION OF THE ACTIVITY OF RENNEN OR 
MILK-CURDLING FERMENT 

Normal gastric juice contains, besides hydrochloric acid 
and pepsin, the rennen ferment as a natural secretory product 
of the gastric mucosa. Eennen possesses the property of coagu- 
lating milk without the presence or assistance of acids. 

Method of Leo. — To 10 cubic centimeters of fresh, 
uncooked neutral or amphoteric milk, add from two to five 
drops of filtered gastric juice, and place the mixture in an 
incubator at 37° C. If rennen is present in normal amount, 
curdling should take place in from ten to fifteen minutes. In 
this process the slight amount of acid contained in the gastric 
filtrate is insufficient to cause coagulation. If curdling takes 
place very slowly it is questionable whether this change is due 
to the action of the rennen or to the formation of lactic acid, 
so to be exact, the reaction of the mixture should be taken 
before and after curdling has occurred. The rennen reaction 
is certain to have occurred only in the presence of an unchanged 
reaction. If coagulation does not occur within an hour rennen 
can be considered absent. As a further guide it may be remem- 
bered that the characteristic curd from rennen is a cake of 



MOTOR-FUNCTION OF THE STOMACH. 137 

casein floating on clear serum, while the curd from lactic acid 
is lumpy and broken. 



DIGESTION OF STARCH AND SUGAR. 

During digestion starch is converted into grape-sugar, while 
cane-sugar is converted into invert-sugar (a mixture of cane- 
and grape-sugar). This action is instituted by the ptyalin of 
the saliva, and continues in the stomach in the presence of low 
acidity. It is later arrested by the increasing acidity, and is 
completed in the intestine by the amylopsin of the pancreatic 
juice. 

Starch is recognized by the deep blue color produced by 
the addition of a dilute solution of iodine or Lugol's solution. 
This reaction grows less vivid as the starch is converted. If 
starch digestion is unduly delayed or does not occur, we may 
infer hyperacidity of the gastric juice. 



DETERMINATION OF THE RATE OF ABSORPTION 
FROM THE STOMACH. 

Penzoldt^s Method. — A capsule containing potassium 
iodide (one and one-half grain) is swallowed. The appearance 
of the iodine reaction in the saliva indicates that absorption has 
occurred from the stomach. To test for the iodide in the saliva, 
paper moistened with starch paste and dried is used. After the 
capsule has been swallowed, the paper is moistened with saliva at 
short regular intervals, and then touched with a glass rod dipped 
in commercial (better fuming) nitric acid. Upon the appear- 
ance of iodine in the saliva the characteristic blue reaction 
occurs. 

When absorption is normal this reaction is usually positive 
in from ten to fifteen minutes ; but if absorption is delayed, the 
reaction may be slow in appearing or occur not at all. 

TEST OF THE MOTOR-FUNCTION OF THE STOMACH. 

If attempted extraction of a full meal six hours after in- 
gestion fails when properly performed, or if nothing can be 



138 THE STOMACH. 

recovered from an Ewald test-breakfast after two and one-half 
hours, the motor function of the stomach may be considered 
normal. 

A second method is to administer a large dose (ten or 
fifteen grains) of salol and test the urine at definite periods for 
the appearance of the products of its decomposition. The com- 
ponents of salol, carbolic and salicylic acid, are separated in the 
alkaline juice of the small intestine. They remain unchanged 
and undissolved during gastric digestion. Salicylic acid is 
readily detected in the urine by the violet color produced by the 
addition of neutral ferric chlorid solution. This test is con- 
veniently performed by moistening filter-paper and bringing a 
drop of the ferric chlorid solution in contact with it. If gas- 
tric peristalsis is normal salicylic acid should begin to appear 
in the urine from forty to sixty or seventy-five minutes after 
ingestion of fifteen grains of salol. 

Iodoform Method. — Give with the test-breakfast one gram 
of iodoform in a well sealed capsule. The iodoform being in- 
soluble in the gastric juice, will not be absorbed by the stomach, 
but will be passed on by peristaltic action to the intestine. By 
the action of the fluids of the duodenum the iodoform is decom- 
posed with the formation of soluble sodium iodide. The demon- 
stration of the iodide in the saliva by means of starch paper and 
nitrous acid, will indicate the time when the gastric contents is 
being discharged into the intestine. Iodine should normally be 
detected in the saliva in from one hour to one hour and a half 
after the ingestion of the capsule. 

ROENTGEN RAY EXAMINATION. 

With the advent of the Roentgen ray and its practical appli- 
cation, it has become an invaluable aid to diagnosis. A number 
of competent men have applied this agent in the study of the 
digestive tract, and have reached so many valuable conclusions 
pertaining to the size, location, motility, etc., of the stomach, 
that the examination of a patient suffering with any disorder 
of the digestive tract must be considered incomplete unless a 
Eoentgenologic examination has been made. 

The following quotation from an article by an expert in 



ROENTGEN KAY EXAMINATION. 



139 



this method of examination and treatment shows the present 
attitude of the profession upon this point 10 : — 

"The superiority of this method of examination over others 
is recognized, I think, by all who have investigated sufficiently, 
and the subject has reached a stage of development when no case 




Fig. 17.— Solid Line shows form and relation of typical normal Stomach, 
with subject in standing posture. dotted llne shows altera- 
TION in position of Stomach, the result of contrac- 
tion of the Abdominal Muscles. 

A, Location of Ensiform. B, Umbilicus. C, Cardia. D, Pylorus. 



involving chronic ailment of the alimentary canal is completely 
investigated until there has been a Eoentgenologic examination 
by a competent man." From a careful examination of a number 
of normal individuals bv the same observer, the following outline 



10 G. E. Pfahler, Jour. Amer. Med. Asso., Dec. 21, 1907. 



140 



THE STOMACH. 



may be considered to be that of a normal stomach in a healthy 
individual : — 

When empty or moderately filled the stomach (Figs. 17, 18 
and Plate II) occupies the left side of the abdomen and extends 
from the inner two-thirds of the left dome of the diaphragm 
to the median line just above the umbilicus. The upper two- 



D 




Fig. 18.— Dotted Line shows form and location of normal Stomach. 

Shaded portion shows contraction and change in form 

of Stomach during filling. 



A, Ensiform. B, Umbilicus. C, Cardia. D, Pylorus. E, Upper Pole. 
Pole. G, Showing place where Peristaltic Waves begin. 



F, Lower 



thirds is almost vertical, and the lower third is almost hori- 
zontal, making the general direction of the stomach slightly 
oblique. In the normal stomach the pylorus is on a level with 
the lower pole. Wo difference has been recognized between the 
normal type in children and in adults. 



PLATE II. 




Normal Stomach. Child, Age 4 Years. 

Instantaneous Postero-anterior Exposure in Standing- Posture. Shows Normal 
Stomach in Normal Position and its Relation to the Heart, Diaphragm, Spinal 
Column and Pelvis. A. Ensiform Cartilage. B. Umbilicus. Arrow Indicates 
Location of Pylorus. {Radiograph by Dr. G. E. Pfahler.) 



ROENTGEN RAY KXAMINATION. 141 

The stomach of the average individual, in the standing 
position, extends to or below the umbilicus. It is vertical for 
more than its upper two-thirds. It occupies the left side of the 
abdomen, except when distended. The pyloric portion extends 
for one to two inches beyond the median line to the right. 

The stomach is normally a very movable organ, as shown 
by the fact that about two-thirds of it crosses the median line 
when the patient lies upon the right side. When distended with 
food the pylorus is carried downward and to the right. 

In summing up the results of his studies of the alimentary 
canal by the Eoentgenologic method, Dr. Pfahler arrives at the 
following conclusions pertaining to its utility in diseases of the 
stomach : — 

"1. The Eoentgen examination will demonstrate obstructive 
disease anywhere along the alimentary canal, and much informa- 
tion concerning its character may be obtained. 

"2. The Eoentgen method is probably the most useful in 
the study of the stomach. By this means the size, form, position, 
motility, effects of massage, respiratory movements, abdominal 
contractions, peristaltic action, and the effect of food can be 
studied. 

"3. The Eoentgen rays are valuable in collecting additional 
data in the diagnosis of carcinoma of the stomach, but must not 
be depended upon to make an absolute diagnosis. 

"4. The Eoentgenoscopic and the Boentgenographic method 
of examination each has its advantage. Eoentgenoscopically 
we study the motion of the viscera, while Eoentgenographically 
we make accurate records and often obtain finer detail." 

Technic of the Method. — Preparation of the Patient: 
The stomach and bowels should be empty, if possible. A purga- 
tive should be given the day before, and if the bowels are not 
moved an enema should precede the examination. Depending 
on the information desired, an ounce of bismuth may be given 
with the full meal, and this mixed with the food by massage 
or muscular contraction, or the bismuth-kefir mixture (bis- ' 
muth subcarbonate, one ounce; kefir, one pint) may be given. 

When carcinoma is suspected, no food should be given be- 
fore the examination because the lodgment of food in the stom- 
ach may give a picture suggestive of neoplasm. Men need no 



142 THE STOMACH. 

special instructions regarding dress. Women should wear a 
kimona or special muslin gown. This special muslin gown 
hangs from the shoulders and should be slit in front at the level 
of the pubes and posteriorly in the lumbar region for the dis- 
tance of ten inches. These slits are closed by strings. Through 
them the necessary land marks are determined, and the cents 
are attached to the ensiform cartilage and the umbilicus by 
adhesive plaster. 

Apparatus. — Besides the usual apparatus for Koentgen- 
ographic examination, appliances are necessary which will enable 
the operator to make the observations and negatives in any posi- 
tion. Eoentgenoscopic apparatus is also necessary, and a dark 
room which is absolutely dark. Dr. Pfahler has recently adapted 
a dark room for this purpose. All the apparatus is in an adjoin- 
ing room. The rays are passed through a door into the dark 
room before reaching the patient. Therefore all light is excluded 
and the patient is not frightened by the noise of the machine, 
nor is there danger of the patient touching the wires. In addi- 
tion this door cuts off the soft rays and make it practically im- 
possible for the skin of the patient to be burnt during the 
observations. 

It is necessary in order to study the shadows on the screen 
that the operator should remain in the dark room for fifteen 
minutes before beginning the examination. 

In order to study both the stomach and the colon at the 
same time, an ounce of bismuth mixture may be given and 
allowed to pass to the colon. After twenty-four hours another 
ounce of bismuth is given, when the outlines of the stomach and 
colon will appear in their relative positions. 



X. 

THE FECES. 



PHYSICAL CHARACTERISTICS. 

The Number. — One stool per day is the normal average for 
a healthy adult. Three daily, or one in forty-eight hours, may 
be normal for some individuals, and not incompatible with 
health. 

The Reaction. — Whether the stools be acid or alkaline is 
of no special clinical importance. In adults the reaction is usu- 
ally alkaline, sometimes neutral, but rarely acid. Acid stools 
are the rule in infants. 

The Amount. — The amount varies in proportion to the 
amount of solids ingested. A preponderance of vegetable food 
usually produces a large quantity, while animal food leaves com- 
paratively little residue. 

The average daily amount of feces varies between 60 and 
250 grams, of which 75 per cent, is water. 

To weigh solid feces ascertain the weight of both the feces 
and their container, then weigh the container empty, and sub- 
tract the latter from the former weight, which will represent 
the weight of the contained feces. If the feces are liquid they 
may be measured, and the amount expressed in cubic centi- 
meters. 

The Odor. — The disagreeable odor is largely due to the 
presence of indol and skatol, but may be further increased by 
hydrogen sulphide, methane, and methyl-mercaptan. 

The Consistence. — This varies greatly and depends largely 
upon the amount of fluids ingested, the temperature, the climate, 
and the condition of the digestive tract. In man the usual form 
is the characteristic plastic cylinder. Clinically expressed, the 
consistence of the feces may be liquid, mushy, or solid or formed. 

Serous Stools. — These are composed of fluid without fecal 
matter, and are of considerable diagnostic importance. Such 

(143) 



144 THE FECES. 

stools are characteristic of Asiatic cholera, cholera morbus, chol- 
era infantum, and poisoning by antimony. In cancer of the 
rectum evacuations are small, frequent, and serous. Arsenic 
poisoning and acute catarrhal enteritis may produce this form 
of stool. Poisoning by toad-stools is also a frequent cause of 
serous stools. 

The Color. — The color varies with the character of the food 
ingested, and is usually little influenced by the decomposition 
products of the biliary pigments. In adults, the color usually 
varies from a light to a dark-brown. Lack of color in the stools 
may occur in health, owing to the over-oxidation of the coloring 
matter into colorless products of bilirubin. Such stools do not 
necessarily indicate the presence of large amounts of fat nor 
obstruction of the biliary passages. 

In infants, fat and undigested milk produce a whitish-color 
tinged with bile-pigments. 

Greex stools are seen after taking calomel. After expo- 
sure to the air such stools are usually acid. 

Yellow stools may be caused by the ingestion of santonin, 
rhubarb, or senna. Typhoid stools are yellowish-brown and have 
received the name of "pea-soup stools/' 

White or clay-colored stools frequently denote the ab- 
sence of bile, as in obstructive jaundice. 

Very dark or black stools may result from the administra- 
tion of iron, manganese or bismuth, or from the ingestion of 
much meat, blackberries or red-wine. 

"Tarry" stools usually denote hemorrhage. 

EXAMINATION OF INTESTINAL DIGESTION BY MEANS 
OF GLUTOID CAPSULES. 

Sahli recommends the use of glutoid capsules which are 
made from gelatin hardened in formaldehyde. These either do 
not dissolve in the gastric juice at all, or only after a consid- 
erable time, but are rather quickly soluble in the normal intes- 
tinal juices. 1 These are preferable to the keratin coated pills 
which were originally recommended, and are useful to diagnose 
the condition of intestinal digestion, e.g., the pancreatic func- 



L Deutch. Med. Woch., No. 1, 1897. 



DETECTION OF IODINE. 145 

tion. They are filled with some substance which does not diffuse 
through the capsule wall, and whose absorption may be studied 
by an examination of the saliva or the urine. For diagnostic 
purposes glutoid capsules, containing two grains (.15 milli- 
gram) of iodoform and four grains (.26 gram) salol, are con- 
venient. 

In order to make the conditions of the test as uniform as 
possible in regard to the length of time the capsules remain in 
the stomach and in regard to the degree of digestive stimulation, 
it is advisable to administer the capsule with the test-breakfast. 
Experience has shown that normally, under the best conditions, 
i.e., normal gastric motility, normal intestinal digestion, and 
normal intestinal absorption, iodine reaction may be expected 
to appear in the saliva, and salicyluric reaction in the urine in 
from four to six hours. 

If one wishes to test the accuracy of any particular lot of 
capsules, it will be necessary to prove that this reaction time is 
obtained in healthy normal individuals. Only rough approxi- 
mate differences in the time of the reaction are of clinical im- 
portance, so that it is sufficient to examine the saliva and the 
urine after six, ten, and twenty-four hours. The best results 
from this method are obtained by administering the capsule in 
the morning upon an empty stomach, and then, four hours later, 
allowing the patient to resume his meals as usual. The speci- 
mens of urine and saliva may, of course, be saved, and exam- 
ined afterward. 



DETECTION OF IODINE IN THE URINE OR SALIVA. 

A few cubic centimeters of the fluid to be examined are 
boiled with a small piece of starch about the size of a pea, until 
the latter is dissolved. After cooling, the mixture is carefully 
overlaid upon nitric acid. If iodine is present a blue- violet ring, 
that gradually disappears, is formed at the line of junction of 
the two fluids. 

A secoxd method is to add to the urine a few drops of 
crude nitric acid and a less number of drops of chloroform, and 
agitate the mixture gently. If iodine be present the chloroform, 
which settles to the bottom, will be tinged rose-red or violet. 



146 THE FECES. 

Both of the above tests are very delicate. But if the urine 
contains a very slight trace of iodine the chloroform-test is not 
very conclusive, since indol, skatol, and urorosein pigments may 
tinge the chloroform red. 



DETECTION OF SALICYLURIC ACID IN THE URINE. 

Dilute ferric chloride is added to the urine drop by drop. 
If the latter becomes a more or less intense violet the reaction 
is positive. 



DETERMINATION OF THE MOTOR FUNCTIONS OF THE 
GASTRO-INTESTINAL TRACT. 

Method of Adolph Schmidt. 2 — Schmidt demands two con- 
ditions for a satisfactory clinical method of examining the feces : 
1. A knowledge of what a normal stool should be under a certain 
diet, involving the use of a "test-diet." 2. The methods of 
examination must be so simple that they are within the reach 
of every physician. 

The Test-Diet. — The requirements are: (a) That it be 
nutritious enough to furnish calories sufficient for the body's 
needs, (b) That it be so constituted that it can be readily ob- 
tained in any household or hospital dietary, (c) That it con- 
tains a known amount of its constituents, so that variations in 
digestion and absorption can be detected in the stool. 

Schmidt suggests the diet which is given below, followed 
by the approximate equivalents as sugested by Steele: — 

Milk, 1.5 liters (2% pints). 

Zwieback, 100 grams (3 ounces well-dried toast). 

Two eggs. 

Butter, 50 grams (1% ounces). 

Beef, very rare or raw, 125 grams (*4 pound). 

Potatoes, 190 grams (6 ounces). 

Gruel made from 60 grams oat-meal (2% ounces). 

Sugar, 20 grams (% ounce). 

This may be given somewhat as follows : — 



■ Quoted by Steele, Medical News, Dec., 1905. 



DETERMINATION OF MOTOR FUNCTIONS. 147 

Breakfast. — Two eggs, half of the amount of toast and 
butter, 2 glasses of milk, oatmeal, and sugar. 

Dinner. — The steak and potatoes, % of the amount of toast 
and butter, 1% glasses of milk. 

Supper. — Two glasses of milk, remainder of toast and 
butter. 

This diet is sufficient for the needs of the average patient, 
and while not offering much variety, is only taken for two or 
three days. A capsule containing five grains of charcoal is given 
with the first meal, and no examination made until the charcoal 
(black) appears in the stool. 

The amount of each article composing the day's dietary 
must first be accurately determined and compared to the con- 
tents of convenient measures, so that the articles need not be 
weighed every meal. This does not apply to the meat and 
potatoes which should be weighed each day. 

The Period of Passage. — The period of time required for 
food to pass through the intestinal tract can easily be deter- 
mined by w r atching for the black in the stool. It is quite as 
necessary to know the time required for the passage of chyme 
through the gastro-intestinal tract, as it is to ascertain how 
often the stools occur, since the two are in no way identical. 

It has been shown in chronic colitis with several watery 
stools a day, that the period of passage may be normal, and that 
peristalsis is decidedly increased only in the colon. According 
to Strass, using a diet similar to Schmidt, the time required for 
food to pass through the alimentary canal varies normally be- 
tween ten and thirty hours. Under abnormal conditions this 
may vary between four and forty-eight hours. 

The Specimen. — After the appearance of the charcoal, 
which indicates the beginning of the passage of the test-diet, 
a portion of one passage is collected in a clean receptacle and 
transferred to a clean, wide-mouthed bottle, and conveyed to the 
laboratory for examination. 

Much can be determined from a naked-eye examination of 
such a mass. The technic for this examination, as outlined by 
Steele, is as follows : — 

Take a piece of formed stool about the size of an English 
walnut or an approximate equivalent of liquid feces, and rub it 



148 THE FECES. 

up in a mortar with distilled water until quite smooth and 
liquid. Part of this is then poured into a Petri dish and 
examined in a good light on a dark background. A little 
experience will enable the observer to acquire a large amount 
of information about the composition of the stool by this 
macroscopic examination. 

The Gross Appearance of Normal Stools. — In normal diges- 
tion nothing should be seen by the naked eye, save a varying 
number of brown points (oatmeal husks), cellulose and indigest- 
ible parts of food, and occasionally sago-like grains that look 
like mucus, but which are found upon microscopic examination 
to be grains of potato. If, on the addition of the water, fat 
globules are seen, they must be considered pathologic unless the 
patient is known to have ingested excessive amounts of fats, as 
olive or castor oil. 

Gross Appearance of Pathologic Stools. — (a) Mucus in 
large or small flakes is not affected by rubbing in the mortar. 
The mucus appears as glassy, translucent flakes often stained 
yellow by bile-pigment. Doubtful cases may be decided by the 
microscope. 

(b) Pus in sufficient quantity to be recognized macro- 
scopically, will have the characteristics of pus in any other 
region. It usually appears as small, yellowish streaks or collec- 
tions throughout the stool. 

(c) Blood, if in appreciable amount, will appear as dark 
reddish-brown or black; if from high in the digestive tube, 
becoming more nearly the color of fresh blood as the source of 
origin approaches the lower end of the bowel. 

(d) Parasites. — Segments of tape-worm, also thread- and 
seat-worms, may be detected in varying numbers in patients so 
afflicted. 

(e) Foreign bodies having entered the digestive tract, if 
indigestible and not impeded in their progress, may be recovered 
from the stool at varying periods after ingestion. 

(f) Calculi from the various glands situated along the di- 
gestive tract, or from the intestinal canal itself (enteroliths), 
may be found in the feces, and appear as calcareous masses of 
various sizes and composition, depending upon their origin and 
age. 



.MICROSCOPIC APPEARANCE OF NORMAL STOOLS. 149 

(g) Remnants of connective tissue and sinew from beef- 
steak. These can be detected by their whitish-yellow color and 
toughness, by which they can be distinguished from mucus. In 
case of doubt the piece should be examined microscopically with 
a drop of acetic acid. Connective tissue then loses its fibrous 
structure, while mucus becomes more thread-like. Small, single 
pieces of connective tissue can be found in normal stools. 

(h) Remnants of Muscle-Fiber. — These appear as small 
reddish-brown threads or small irregular lumps. 

(i) Remnants of Potato. — These appear like boiled grains 
of tapioca and may easily be confused w r ith mucus. The micro- 
scope will show the true nature of these bodies. 

(j) Large crystals of triple phosphate occur in foul stools, 
and can be recognized by their shape and by their solubility in 
all acids. 

Microscopic Appearance of Normal Stools. 3 — Three slides 
are prepared from the liquid feces. The first is merely a drop 
of the material to be examined by both low and high power. 
The second slide is prepared by mixing a drop of acetic acid and 
a drop of the liquid feces upon a slide, heating it to boiling and 
then putting on a cover-glass. The third slide is prepared by 
mixing a drop of liquid feces with a drop of Lugol's solution. 

Microscopic Examination of Slides. — Slide 1 will 
show (a) single small muscle fibers colored yellow with cross 
striation. Visible with a Leitz 3, but showing better with higher 
power, (b) Small and large yellow crystals of salts of the fatty 
acids, (c) Colorless (gray) particles of soap, (d) Single potato 
cells without distinguishable contents, (e) Particles of oat- 
meal and grain husks. 

Slide 2. — A general idea of the fat-content of the stool may 
be obtained. Upon cooling, small drops of fatty acids may be 
found covering the whole preparation. The large crystals of 
salts of the fatty acids are broken up by the acetic acid, and 
fatty acids liberated. If the slide is heated again and exam- 
ined hot, the fatty acids will be found to run together in drops 
which, as the slide cools, break suddenly apart. 



3 Technic of E. Dutton Steele, Medical News, Dec. 16, 1905. 



150 THE FECES. 

Slide 3. — Here should be found violet-blue grains in some 
of the potato cells, and small single blue points, probably fungi 
or spores. 

Pathologic Microscopic Findings. — Slide 1. (a) Muscle 
fibers in excess, perhaps with yellow nuclei, (b) Xeutral fat 
drops, or fatty acid, in soap crystals, (c) An excess of potato 
cells with more or less well-preserved contents, (d) Parasite 
eggs, mucus, connective tissue, pus, etc. 

Slide 2. — Fatty-acid droplets in excess. 

Slide 3. — Blue starch grains in the potato cells or free, oat- 
meal cells, fungi, spores or mycelia. 

Formed Elements in the Feces. — (a) Blood. Eed cells, 
if recently shed, may be distinguished by their characteristic 
form, or if disintegrated, only masses of brownish-red amorphous 
hematoidin will be found. In a certain percentage of cases char- 
acteristic crystals of hematoidin will be found. 

(b) Epithelial Cells. — These are normally present in mod- 
erate numbers and represent the natural desquamation from the 
intestinal canal. They are more or less disintegrated, depending 
to some extent upon their height of origin in the intestinal canal, 
and upon the length of time they have remained free in the 
digestive tube. In catarrhal conditions they may be present in 
very large numbers, when they may assume diagnostic impor- 
tance. 

(c) Pus Cells. — These rapidly undergo decomposition, so 
that even when numerous and coming from a comparatively 
short distance above the rectum, they may be beyond recogni- 
tion. The characteristic pus cell appears as a small round or 
slightly oval granular body. The presence of very much pus in 
the stools is indicative of rupture of an abscess into the intes- 
tinal tract. 

CHEMICAL EXAMINATION. 

This comprises only five routine tests: 1. The reaction. 
2. The sublimate test for the condition of the bile salts. 3. The 
fermentation test. 4. The test for "lost albumin." 5. The test 
for occult blood. 

1. The Reaction. — This is quite difficult to get with the 
ordinary litmus paper. It can be easily determined by dropping 



CHEMICAL EXAMINATION. 



151 




Fig. 19.— Strasburger Apparatus, showing arrangement of Bottles 
for Fermentation Test of Feces. (After Steele.) 



152 THE FECES. 

a little softened fecal matter into 5 or 10 cubic centimeters of 
weak watery solution of neutral litmus, shaking it and noticing 
the color reaction. 

2. The Sublimate Test. — This consists of taking a few 
cubic centimeters of liquid feces and mixing with an equal 
amount of a saturated watery solution of HgCl 2 . A normal 
stool will quickly turn a pinkish-red, indicating the presence of 
hydrobilirubin, which will be more intense the fresher the 
material. A green color is pathologic and indicates the presence 
of unchanged bile-pigment. 

3. Fermextatiox Test. — Described by Steele, using his 
modification of Strasburger's apparatus. 

The apparatus consists of a two-ounce, wide-mouth bottle 
A (see Fig. 19). This is fitted with a perforated cork through 
which runs a tube to the test-tube B, which is also fitted with 
a rubber cork with two perforations. A bent tube runs from 
the tube B to the test-tube C, to allow for the escape of air. 
Each tube has a capacity of a little more than 30 cubic centi- 
meters when fitted on the corks. The apparatus is simple and 
easily constructed, and broken parts can be replaced readily. 

The Test. — About five grams of solid feces, or an equivalent 
of liquid feces, are rubbed up with a little distilled water and 
placed in the main bottle A. This is filled with sterile water, 
the tube is filled with water and fitted into place (not neces- 
sarily full), and tube C is then fitted on empty. The reaction 
is carefully noted before the test is started. The apparatus is 
then stood in a warm place — best in an incubator at 37° C. for 
twenty-four hours. If gas forms by fermentation in A it will 
rise into B, and the amount will be indicated by the amount of 
water displaced into C. formally the fermentation test should 
show practically no gas, and the original reaction of the material 
should be unaltered in twenty-four hours. If more than one- 
third of the tube C is filled, it is pathologic. If the reaction 
after twenty-four hours is decidedly more acid, it is a carbo- 
hydrate fermentation. If alkaline, and with a foul smell, it is a 
fermentation of the albumins. 

4. Test for Lost Albumin. — A portion of softened stool 
is filtered (a slow and difficult process). The filtrate is shaken 
with silicon and refiltered, then is saturated with acetic acid to 



BLOOD IN THE STOOL. 153 

bring down the nucleo-proteids, and finally a drop of potassium 
ferrocyanide solution is added. A decided precipitate indicates 
albumin. A positive test shows only that there is a decided 
diminution in albumin digestion. 

BLOOD IN THE STOOL. 

If bleeding has its origin in the upper part of the digestive 
tract (stomach or small intestine) , it is so altered by the action 
of the digestive fluid that, by the time it finally appears in 
the stool, it has a black or brownish-black appearance which has 
been likened to tar or coffee grounds. However, if the hemor- 
rhage be large and the peristalsis very active, blood may appear 
almost unchanged in the stool, even when of high origin in the 
digestive tube. 

Eelatively small amounts of blood may be so changed and 
mixed with the feces that they cannot be detected by the naked 
eye or by the microscope. This is termed "occult" blood, and 
is only recognizable by chemical means. 

Preliminary Technic. — Owing to the possibility of the pres- 
ence of a positive reaction, resulting from the ingestion of hemo- 
globin-containing food in a normal individual, it becomes neces- 
sary to restrict the diet in order to eliminate this possible source 
of error, and to limit the test-diet by the administration of a 
capsule containing five or ten grains of charcoal, and to watch 
the stools for the appearance of the black discoloration due to 
the passage of the charcoal. Only after the appearance of the 
charcoal in the feces should a specimen be taken for the test for 
occult blood. 

Steele 4 recommends a "liquid diet/' including milk and 
broths, or a "semi-liquid diet" composed of milk, eggs, and toast, 
to which may be added moderate amounts of the ordinary winter 
vegetables. 

Eed meats and beef juice should positively be withheld. 
Steele has shown that iron, either in the organic or inorganic 
form, will even in large doses not affect the reaction. 

As a preliminary to the test, it is necessary to eliminate as 
fully as possible extraneous sources of blood. Thus tuber- 



4 Amer. Jour. Med. Sci., July, 1905 



154 THE FECES. 

culous ulcer, typhoid fever, hemorrhoids, fissure, and pur- 
pura should be excluded. Also ingestion of carmine, swallowed 
blood from any cause, hemoptysis, epistaxis, and menstruation. 

Technic of Test. 5 — If the feces are solid they must be 
softened with distilled water. To 5 or 6 cubic centimeters of 
liquid feces, in a wide-mouthed cork-stoppered bottle, add about 
thrice as much ether and agitate or thoroughly mix by shaking. 
Then add a few grains of powdered guaiac and again agitate. 
Follow this by 5 cubic centimeters of glacial acetic acid (99.4 
per cent.), and still again agitate. Allow this mixture to stand 
until the solid particles settle to the bottom, and then decant into 
each of two test-tubes 5 cubic centimeters of the supernatant 
liquid. 

One test-tube should be kept as a control. To the other add 
1 or 2 cubic centimeters of a fresh solution of hydrogen dioxid. 
If a bluish discoloration occurs either at the line of contact of 
the two solutions or throughout the mixture, the reaction is 
positive. 

Significance of the Occult Blood. — According to Boas 
occult blood is constantly present in carcinoma of the gastro- 
intestinal tract; intermittently present in gastric and duodenal 
ulcers; occasionally in stenosis of the pylorus, and is absent in 
gastritis, hyperchlorhydria, and in the gastric neuroses. 

BACTERIA AND PROTOZOA IN THE FECES. 

There are many varieties of bacteria in the feces. They 
have been estimated to amount to about one-third of dried feces. 
Some are harmless at all times; others which are, under ordi- 
nary conditions, harmless may develop pathogenicity under cer- 
tain circumstances. 

Bacillus Coli Communis. — This organism is constantly pres- 
ent in the feces, and is normally non-pathogenic. It has, how- 
ever, been found in pure culture in cases of appendicitis, 
empyema of the gall-bladder, pyelitis, and cystitis. 

Morphology. 6 — It is a rod with rounded ends, sometimes 
so short as to appear almost spherical, while again it is seen with 



5 From Professor Daland's Laboratory. 

6 Abbott's Bacteriology. 



BACTERIA AND PROTOZOA IN THE FECES. 155 

very much longer threads. It may occur as single cells or joined 
together in pairs, end to end. It is motile, and does not form 
spores. It stains with the ordinary alkaline dyes, and is decol- 
orized by Gram's method. 

Compared to the typhoid bacillus, it will be found to be 
less motile; grows more rapidly on gelatine colonies, and lux- 
uriantly on potato (typhoid growth usually invisible). Colon 
bacillus coagulates milk in incubator from thirty-six to seventy- 
two hours; typhoid does not. Colon bacillus decomposes sugar 
solutions; typhoid does not. 7 

Bacillus Typhosis. — This is a distinctly pathologic bacte- 
rium, and is the specific cause of typhoid fever. Its appearance 
in the stools is scanty, but usually can be isolated by appropriate 
methods during the first few days of the disease. 

Pratt, Peabody, and Long 8 obtained it from the stools of 
only 31 per cent, of febrile cases, and none in twenty-one con- 
valescents examined by them. They state that it occurs most 
in the blood, and that it does not develop in the intestinal con- 
tents except under unusual conditions. The bacillus in the in- 
testines comes chiefly from the gall-bladder; frequently it is 
found in the urine in larger numbers than in the feces, and is 
found in greatest numbers in feces containing blood. 

Morphology. — The bacillus is about three times as long 
as it is broad, with rounded ends. Its length may vary greatly, 
but its width remains fairly constant. It is actively motile, and 
Loeffler's method of staining will show it is possessed of many 
delicate flagelli (see methods of staining). 

Its growth on potato usually is invisible. It does not cause 
coagulation of milk (colon bacillus does). Owing to the tend- 
ency to retraction of the protoplasm from the cell-envelope, 
and the consequent production of vacuoles in the bacilli, the 
staining is often irregular. It stains with the ordinary aniline 
dyes. 

Bacillus lactis aerogenes and the bacillus proteus vulgaris, 
are probably always present in the stools. They are probably 
pathologic in many cases of cholera infantum. 



7 For methods of plating, isolating, and culture, see works on bacteriology. 
8 Jour. Amer. Med. Asso., Sept. 7, 1907. 



156 THE FECES. 

Streptococcus aerogenes. — This is an etiologic factor in 
certain cases of entero-colitis. 

Bacillus Dysentericus (Shiga bacillus) is the cause of one 
form of infantile dysentery, and is generally present in the dis- 
charges of such patients showing blood and mucus. 

The Comma Bacillus. — This is the infective agent in Asiatic 
or true cholera. 

Morphology. — It is a slightly curved rod of a length of 
from one-half to two-thirds that of the tubercle bacillus, but 
thicker. Its curve is never very marked, it may be nearly 
straight. It is a flagellated organism, but has only one flagellum 
attached to one end. It is actively motile. It does not form 
spores. 

Cultural Characteristics. — On gelatine plates at room- 
temperature, its development can be observed after as short a 
period as twelve hours. It rapidly liquefies gelatine. It is 
strictly aerobic. It takes the ordinary stains. 

Diagnostic Method. — A smear is made from one of the 
small, slimy particles found in the semi-fluid evacuations, dried, 
fixed, and stained in the ordinary way. If upon microscopic 
examination only curved rods, or curved rods greatly in excess of 
all other organisms, are found, the diagnosis of Asiatic cholera 
most probably is correct. 

Tubercle Bacillus. — This organism probably is always pres- 
ent in all cases of intestinal tuberculosis. 

Diagnostic Techxic (E. C. Bosenberger's). — A selection 
of "any part of feces is made, there being no effort at a selection 
of any particular mass or portion. If the stool is solid a small 
mass is mixed with sterile water. After drying and fixing, the 
spread is stained with carbol fuchsin for fifteen or twenty 
minutes, cold. The excess of stain is drained off and Pappen- 
heim's solution (see Appendix) is applied, and when the 
preparation is the color of the counter stain (methylene-blue) 
thorough washing in distilled water is resorted to, the spread 
being then dried and mounted in balsam. The most important 
point in the technic is to obtain a spread the color of the counter 
stain with not a particle of the carbol-fuchsin showing to the 
naked eye." 9 



9 Solis-Cohen : New York Med. Jour., Aug. 21, 1907. 



CLINICAL SIGNIFICANCE OF EXAMINATIONS. 157 

It is alleged, but not conclusively determined, that no other 
acid- or alcohol-fast bacilli will withstand this method except- 
ing the tubercle bacillus. 

Ameba Dysenteriae. — This is now generally conceded to be 
the specific cause of the so-called amebic dysentery. In typical 
cases the stools contain much blood-stained mucus, containing 
large numbers of these amebas. These parasites so resemble 
epithelial cells that a positive diagnosis can only be reached when 
they are seen under the microscope to move and to extend their 
psendopods. To keep them alive sufficiently long for examina- 
tion, the feces must be caught in a warm pan and a portion im- 
mediately transferred to a warm microscope slide for examina- 
tion. The organism is from 8 to 50 microns in diameter (see 
Plate I). To develop the nucleus the organisms should be 
obtained fresh; they are killed by the addition of a few drops 
of acetic acid or corrosive sublimate (sat. watery solution) while 
on the microscope stage. The nucleus is spherical and about 5 
microns in diameter (see page 103). 

For intestinal worms, ova, etc., see section on "Animal 
Parasites." 



THE CLINICAL SIGNIFICANCE OF THE EXAMINATIONS. 

Mucus. — As a rule, the appearance of mucus in the stool 
indicates the presence of inflammation of the mucous membrane, 
and is the one trustworthy sign of that condition (Steele). 
There are two conditions in which mucus has no significance: 

1. When thin mucus spreads over the surface of a hard stool. 

2. The so-called mucus-colitis with the discharge of mucus casts. 

Bile-Pigment. — A green color of part or all the stool (by 
the sublimate test) is pathologic, except in children. It means 
too short a period of passage through the intestines. A fresh 
normal stool should give a pink reaction with HgClo. If a color 
reaction of any kind is absent it indicates either absence of 
bile from the intestine, a very fatty stool, or the reduction of 
hydrobilirubin (urobilin) into leucohydrobilirubin, a colorless 
product (Steele). 

Fat. — It will require some practice in the use of the diet to 
tell whether there is an excess of fat in the stool or not. As the 



158 THE FECES. 

normal amount of fat varies within wide limits in normal feces, 
only a great excess can be considered abnormal. 

Remnants of Meat. — ^Normally there should be only micro- 
scopic particles of connective tissue and muscle fiber. An excess 
of either is often visible to the naked eye. 

Excess of connective tissue means insufficient gastric diges- 
tion, because fibrous tissue is digested in the juices of the stom- 
ach only. Excess of undigested muscle-fiber means disturbance 
in intestinal digestion, and probably means trouble in the upper 
part of the small intestine, as follows: (a) Indicating a total 
lesion of the pancreas and absence of proteolytic and tryptic 
digestion, (b) Trypsin may be present, but its activating en- 
zyme, enterokinase, may be absent, (c) The period of passage 
of the food through the intestine may be so rapid that no time 
is given for its digestion, (d) Large pieces of undigested mus- 
cle may be present, because gastric digestion is imperfect and 
the connective-tissue framework of the meat has not been prop- 
erly removed. 

Pathologic carbohydrate fermentation means poor starch 
digestion and indicates, as a rule, disturbance in the small intes- 
tine, which is usually due to insufficiency in the succus entericus. 

Pathologic albumin fermentation means a large remainder 
of albumin in the feces, and indicates, in Schmidt's experience, 
serious trouble, usually some anatomic change in the mucous 
membrane of the small intestine (Steele). 



FOREIGN BODIES, CALCULI, AND CONCRETIONS. 

An attack of biliary colic or gall-stones may be followed by 
the appearance, from time to time, of gall-stones in the feces, 
or they may appear without the previous occurrence of any 
symptoms referable to the liver or its appendages. In search- 
ing for them, the feces should be rendered thoroughly fluid by 
rubbing in water and then by passage through a sieve. Various 
forms of simple and complicated apparatus are upon the market, 
and are known as stool-sieves. A very satisfactory home-made 
substitute is made by filling in a wire-frame with two or three 
layers of gauze or cheese-cloth, the wire-frame being arranged to 
fit around the rim of the bowl of the closet. To be certain of 



FOREIGN BODIES, CALCULI, AND CONCRETIONS. 159 

not missing the stones it is necessary to examine all movements 
for at least fourteen days following the cessation of the attack 
of colic. Even a careful and prolonged search may fail to find 
the concretion in certain undoubted cases of gall-stone disease. 
Sometimes because the stone causing the symptoms was not 
passed, but after being jammed in the neck of the bladder finally 
returned into the cavity of the gall-bladder itself. Again, the 
stone may be retained for a long time in a fold or diverticulum 
of the intestinal tract, and finally the concretion may have fallen 
to pieces in the intestine. Another explanation of why the stones 
fail to appear is that typical attacks of colic may be due to in- 
flammation without the presence of gall-stones. 

Gall-stones are concretions which form in the biliary pas- 
sages. They vary in size from a pin-head to that of a pigeon's 
egg or even larger. They are composed chiefly of cholesterin 
and calcium-bilirubin in varying proportions; besides these 
there are present minute amounts of other bile-oxidation prod- 
ucts and calcium-carbonate. A predominance of cholesterin 
produces a light-, and of calcium-bilirubin a dark-colored stone, 
the absolute color varying usually between a light- or dark-brown 
to a dark-olive green. Stones vary greatly in hardness, and on 
cross-section usually show distinct concentric layers of crys- 
talline substance, sometimes of different colors. The sur- 
faces may present a beautiful faceted formation from attrition 
between a number of stones lying in the bladder, or they may 
be irregularly granular. 

It is important not to confound other intestinal concretions 
with gall-tones. Woody bits of food, particularly the cores of 
pears, have been termed pseudogall-stones. The microscopic 
examination of bits of these scraped off with a knife, present the 
picture of characteristic wood cells. Chemical examination will 
also prevent mistake (see below). 

So-called biliary sand, in most instances, consists of these 
small pseudogall-stones. The existence of true biliary sand has 
not yet been conclusively proven. Another kind of pseudogall- 
stone consists of balls of fat and fatty soaps which are not easily 
melted. They are found in the stools after the administration 
of large amounts of olive oil, as in a favorite method of treating 
cholelithiasis. 



160 THE FECES. 

For chemical examination gall-stones are at first dried, then 
powdered and treated with alcoholic ether to extract the choles- 
terin. This may then easily be recognized by allowing the 
extract to evaporate upon a w T atch-glass, when the characteristic 
glistening rhomboids of cholesterin crystallize out, and may 
be easily recognized by the microscope. After extraction with 
alcohol and ether the residue is treated, while cold, with very 
dilute potassium hydroxid. If the powder contains calcium- 
bilirubin, a yellow solution will be obtained which gives Gmel- 
lin's reaction. 

The much rarer pancreatic concretions differ from gall- 
stones in that they contain no bile-coloring matter, and are com- 
posed chiefly of calcium-carbonate, which dissolves readily with 
effervescence in hydrochloric acid. 

Intestinal Stones or Fecal Concretions. — These are sup- 
posed to play an important part in exciting attacks of appendi- 
citis, but seldom appear in the stools. They consist almost 
exclusively of ammonio-magnesium phosphate (triple phos- 
phate), and should be examined after the manner of urinary 
calculi (see page 22 6 ). 10 



3 Sahli's Diagnosis. 



XL 

THE URINE. 
PART I. 

GENERAL CONSIDERATIONS. 

The normal constituents of the urine are usually tested for 
by quantitative methods, since we are concerned with the actual 
amount of these substances and not with their presence or ab- 
sence from a given sample. In testing for abnormal constitu- 
ents, on the other hand, we are concerned, as a rule, with their 
presence or absence, though in certain instances we may desire 
to know the absolute quantity of these abnormal substances. 

The Sample. — If we are to make a qualitative examination 
for abnormal constituents in a single sample of urine, it is best 
to collect a specimen about three hours after the ingestion of 
a hearty meal (dinner), as such a sample is most likely to con- 
tain the substances sought (usually albumin or sugar). 

Cases which at times show a trace of albumin will usually 
show an increase late in the day or after active exercise, and the 
collection of specimens should be timed to meet these conditions. 

Method of Collection of Specimens. — The following is 
a copy of the directions furnished by Prof. Judson Daland for 
the instruction of his patients : — 

Directions for Collecting Urine. — The urine should be collected 
in a perfectly clean vessel and four ounces sent to the laboratory. Wide- 
mouthed four-ounce bottles especially adapted for this purpose should be 
obtained at the drug stores, and when possible the urine should be 
directly passed into the bottle. 

1. The evening specimen is to be obtained in the following manner. 
Empty the bladder immediately before the evening meal and discard this 
urine. From the urine first passed after the evening meal, take four 
ounces and note the hour when voided. 

2. The second specimen is obtained from the urine first passed upon 
arising in the morning. Note the hour when the urine was passed. 

To obtain the total quantity of urine passed in 24 hours. On the 
day when the observation is begun, at a definite hour, empty the bladder, 

11 (161) 



162 THE URINE. 

and discard this urine. All the urine passed afterwards is to be collected 
in a suitable, clean, dust-proof receptacle and kept in a cool place. The 
following day at the same hour, when the bladder was first emptied and 
the urine discarded, again empty the bladder. This urine should be 
added to complete the total amount for 24 hours, which should be ex- 
pressed in ounces. After the total amount of urine has been collected 
and thoroughly mixed, send four ounces of the mixture. 

Example. Observation began on January 1st., at 8 A.M. The blad- 
der is emptied at 8 A.M., this urine is discarded; the urine passed dur- 
ing the day and night are saved. The next morning, January 2nd., at 
8 A.M., the bladder is again emptied, and this urine is added to complete 
the total quantity for 24 hours. A label on which is written the name, 
date, and time when the urine is passed should be pasted on the bottle. 

The Twenty-four Hour Specimen. — For accurate results, 
particularly by quantitative methods, it is necessary to examine 
a portion of the mixed urine voided during twenty-four hours. 
Such a specimen should be examined within six or twelve hours 
after the collection is complete; this will usually prevent errors 
due to the processes of decomposition and putrefaction which 
might destroy the formed elements. 

Catheterized Specimen. — Catheterization is often resorted 
to in order to obtain urine free from contamination, which might 
enter it from the lower part of the urinary tract. For specimens 
of urine from one kidney the Harris segregator may be used, or 
in the cases of women, the ureters may be catheterized. 

Urine kept at room-temperature, particularly in summer 
time, readily undergoes decomposition and putrefaction. These 
changes render it unfit for examination. 

DECOMPOSITION CHANGES IN NORMAL URINE. 

If no method of preservation of the sample is employed, 
the fresh urine, which is clear of acid reaction and showing no 
deposit, will gradually undergo the following changes: — 

1. The lower part grows cloudy from sedimentation of 
mucus, cells and other detritis, the urine is still acid. 

2. This sediment gradually settles to the bottom and may 
show minute crystals of uric acid, the urine is less acid. 

3. Uniformly cloudy from beginning, precipitation of phos- 
phates. The urine is very faintly acid or neutral. 

4. Very turbid from precipitation of phosphates and devel- 
opment of bacteria. Copious sediment of triple and amorphous 
phosphates, bacteria, ammonium urate, and epithelial debris. 
Alkaline reaction and ammoniacal odor. 



DESCRIPTION AND IMPORTANCE. 163 

PRESERVATION OF SAMPLE. 

When, for any reason, it becomes necessary to delay the 
examination of urine past the time when decomposition changes 
usually occur, these changes may be retarded in a number of 
ways: 1. By refrigeration. 2. By the addition of two or three 
grains of chloral for each ounce of urine. 3. By the addition of 
ten drops of 4-per-cent. formaldehyde solution for each ounce 
of urine, -i. By shaking with chloroform in the amount of five 
drops to the ounce of urine. 

DESCRIPTION AND IMPORTANCE OF THE URINE. 

The urine is an aqueous solution of the by-products of 
metabolism, so far as these are not excreted or eliminated by 
the lungs, bowels, or skin. It is the most important excretory 
product of the body, and is the medium through which the end- 
products of nitrogenous metabolism and the soluble mineral salts 
are almost exclusively excreted under normal conditions. Ab- 
normal products of metabolism and many substances which have 
found their way into the circulation from without and w r hich 
are foreign to the body, are likewise carried out in solution. 

Not less than 50 per cent, of the total fluid ingested daily 
is excreted as urine. 

General Characteristics. — Xormal urine is perfectly trans- 
parent when voided, but soon becomes turbid, and on standing 
deposits a light flocculent sediment composed of a mucinous body 
and a few epithelial cells and leucocytes. If the urine is kept 
cold and care is exercised to exclude the entrance of micro-organ- 
isms, the upper portion of the urine will remain clear indefin- 
itely. Ammoniacal fermentation, due to the activity of the bac- 
terium area and the micrococcus urea, causes cloudy urine from 
the precipitation of the phosphates. 

Bacteria-free urine may, in winter, become cloudy because 
the contained urates are less soluble in cold urine than in warm 
urine. On standing the urates settle to the bottom, and the 
supernatant liquid remains clear as long as bacterial contamina- 
tion does not occur. 

Passage of Turbid Urine. — In man the passage of turbid 
urine is always abnormal, except during the first days of life, 



164 THE URINE. 

when the turbidity is due to the profuse desquamation of epithe- 
lial cells and the relatively large amount of urates. 



PHYSICAL CHARACTERISTICS OF THE URINE. 

The Color. — The color of urine normally varies from a light 
yellow to a dark amber. This is largely influenced by the con- 
centration of the secretion and by the reaction. The pigmenta- 
tion is due chiefly to the presence of a substance called uro- 
chrome (derived from the biliary pigments) and indoxyl potas- 
sium sulphate (indican). Acid urine is always darker than alka- 
line; the color is naturally lighter when the excretion is 
abundant than when it is scant. 

Deviations from the normal color are notably observed 
in certain diseases and during the administration of certain 
drugs. It may also occur in apparently healthy individuals in 
consequence of certain undetermined anomalies of metabolism. 

Pale urine may occur as a neurosis or during the course of 
certain nervous diseases, particularly in epilepsy and hysteria. 
It may be a symptom of chronic nephritis and diabetes. 

Dark urine, which is clear, occurs in the course of most 
acute fevers, and is due to the presence of uro-erythrin. A 
smoky color denotes the presence of decomposed blood, as in 
acute nephritis. Blood-red or pink urine usually denotes the 
presence of fresh blood. Eecently Bar and Duaney 1 have called 
attention to a false bloody urine due to the activities of a pseudo- 
membranous and chromogenic bacterium. 

Yellow-brown or greenish urine suggests the presence of 
bile. Brownish urine occurs in melanosis and after the inges- 
tion of rhubarb, senna, or tannic acid. 

Smoky-brown urine indicates the presence of the end-prod- 
ucts of ingested carbolic acid or its analogues. . 

Pale greenish urine, with a high specific gravity, usually 
indicates glucose. 

White urine denotes the presence of pus or chyle. 

Whitish turbidity denotes: 1. Pus. 2. Phosphates. 3. Am- 
monium urate. 



1 Le Progfres Medicale, Mar. 24, 1906. 



PHYSICAL CHARACTERISTICS. 165 

Urine which Darkens on Standing. — Some urine, after 
standing for a variable period, becomes dark-brown or black. 
This may be due to the presence in the specimen of melanin or 
of phenol. Non-pathologic phenol urine or "carbol urine" may 
arise from medicinal or surgical treatment with phenol or closely 
allied compounds. Under these conditions the end-products in 
the urine usually appear as resorcin. Such urine, if alkaline 
when voided, rapidly darkens on exposure to the air, and if acid 
becomes dark more slowly, as the alkaline reaction gradually 
develops. 

Under certain pathologic conditions the urine may darken 
on exposure to the air, owing to the presence of alkapton. This 
condition is known as alkaptonuria. A similar darkening in 
pathologic urine may result from the presence of a coloring 
matter termed melanogen. 

Test. — To differentiate between the phenols and melan- 
ogen: Add bromine w r ater to the suspected urine. In the 
presence of phenols there will be produced a permanent, yel- 
low precipitate. A primary yellow precipitate which gradually 
darkens, becoming finally brown or black, indicates melanogen. 

Corroborative Test. — Add to the fresh undarkened urine a 
few drops of dilute solution of neutral ferric chloride. A violet 
discoloration denotes phenols, brown or black denotes melan- 
ogen. 

It must be remembered that urine containing alkapton will 
give many of the chemical tests for sugar which depend on a 
reduction process. 

The Odor. — Freshly passed normal urine has a peculiar 
characteristic aromatic odor, resulting from the contained 
volatile acids. Decomposing urine has the characteristic odor 
due in part to the free ammonia resulting from the decomposi- 
tion of urea. 

Urine which is ammoniacal when freshly passed, points to 
a pathologic fermentation occurring in the bladder, usually ac- 
companying cystitis. 

A putrid odor denotes putrefactive change occurring in pus 
or other albuminous substances. 

An odor resembling acetone is occasionally observed and 
denotes diabetes mellitus. 



166 THE URINE. 

Urine containing cystin may, upon standing, develop the 

Odor of HYDROGEX SULPHIDE. 

Some articles of food, as asparagus and onions, and certain 
aromatic medicines, as turpentine and copaiba, may give char- 
acteristic odors. 

THE AMOUNT. 

The average daily excretion of urine in the United States 
is somewhat less than in foreign beer-drinking countries. A 
fairly normal average for the United States may be stated to be 
between 1200 and 1600 cubic centimeters, or about fifty ounces 
for men, while for women it is slightly less. Generally speaking, 
the amount will vary in inverse ratio to the insensible perspira- 
tion: hot weather diminishes and cold weather increases the 
amount. Profuse perspiration, sweating, vomiting, and diar- 
rhea all decidedly diminish the amount. Children, and espe- 
cially nursing infants, on account of the preponderance of liquid 
in their food, excrete a proportionately larger amount of urine, 
compared with body-weight, than do adults. 

The normal amount of urine passed in twenty-four hours is 
consequently subject to wide variations, depending on the amount 
of fluid ingested, the character and the quantity of the food, 
the process of digestion, the blood-pressure, the surrounding 
temperature, the emotions, sleep, exercise, age, sex, and body- 
weight. During repose much less urine is excreted than during 
activity, hence the excretion during the night is less than during 
the day. The maximum secretion is usually observed during the 
first few hours succeeding a hearty meal. 

Artificially the excretion may be increased by substances 
which have a tendency to raise blood-pressure, as tea, coffee, 
and alcohol. Many drugs bring about the same result. The 
most important of the medicinal diuretics are digitalis, squill, 
broom, juniper, nitrous ether, urea, etc. Distilled water also 
possesses distinct diuretic properties. 

The quantity of urine in pathologic conditions depends, first 
on the condition of the secreting renal parenchyma and, sec- 
ondly, upon the condition of the blood-current in the kidneys. 
It will therefore be affected in general by circulatory disturb- 
ances, as well as by disease of the kidney itself. To appre- 



VARIATIONS IN THE AMOUNT. 1(57 

eiably alter the amount of the excretion, both kidneys must be 
diseased, for if one kidney be healthy it will assume vicariously 
the function of the other organ. This compensatory action 
regularly occurs after extirpation of one kidney. 

As a rule the more acute the nephritis the more the excre- 
tion of urine will fall below normal, while the more chronic the 
process the more will the amount exceed normal. This increase 
reaches its maximum in the true contracted kidney where the 
volume excreted in twenty-four hours may be very great. Dis- 
eases of the heart and lungs, leading to chronic passive conges- 
tion, will diminish the total amount of the urine, a condition 
evidently dependent upon interference in the renal circulation. 

It is evident, therefore, that the determination of the total 
output of the kidneys and an observation of the variation in the 
amount during the course of disease, may be of great diagnostic 
and prognostic value. 

The increased excretion of urine following convulsions, par- 
ticularly the hysterical variety and after attacks of angina pec- 
toris, is probably dependent upon some vasomotor disturbance. 
Many alterations in the volume of the urine depend upon quan- 
titative and qualitative variations in the substances eliminated, 
and are primarily induced by disturbances in metabolism. 

Anuria, or entire failure to void urine, may be due either 
to complete suppression of the secretion in the kidneys or to 
obstructions in the urinary tract. 

Hydruria. — This is a state of the urine in which the fluid 
is increased out of proportion to the solids, and is usually asso- 
ciated with an increase in the total twenty-four hours' output. 

Polyuria. — This term is applied to an increase in the elimi- 
nation of the urine as a whole, both fluids and solids. 

Oliguria is applied to a diminution in the total excretion 
of the urine. 

Polyuria may be noted in the following conditions: 1. Dia- 
betes mellitus. 2. Diabetes insipidis. 3. Chronic interstitial 
nephritis. 4. Amyloid disease of the kidney. 

Oliguria is met under the following circumstances: 1. 
Valvular heart-disease. 2. Diminished blood-pressure (see sec- 
tion on blood-pressure). 3. Acute articular rheumatism. 4. 
Chronic parenchymatous nephritis. 5. Acute congestion and 



168 THE URINE. 

inflammation of the kidneys. 6. Decreased cell activity in 
shock. 7. Failure of nutrition preceding death. 



SPECIFIC GRAVITY. 

The average specific gravity of normal urine of 1500 centi- 
meters (fifty ounces) volume for twenty-four hours, is about 
1020. Slight variations (1015 to 1028) from this standard are 
consistent with perfect health and depend chiefly u,pon the char- 
acter of the food ingested, the quantity of water taken, and upon 
the state of metabolism. Under certain conditions of apparent 
health, as after a hearty meal, the specific gravity of the indi- 
vidual specimen may be as high as 1035, and after excessive in- 
gestion of fluids it may temporarily fall to 1005. 

Under pathologic conditions the specific gravity may vary 
between 1001 and 1055 or even higher. If the diet consists 
largely of nitrogenous food it will furnish a relatively larger 
amount of solids; in consequence the specific gravity of the 
urine will be increased. Active muscular exertion also tends to 
raise the specific gravity by increasing tissue catabolism. Co- 
pious diaphoresis may bring about a concentration of the urine 
by diminishing the amount. Fasting has a similar effect. 

The specific gravity usually varies inversely with the volume 
both under normal and pathologic condition. A scanty urine is 
more concentrated than a profuse one. Diabetes mellitus forms 
an exception to this rule, since the presence of sugar produces 
a high specific gravity in spite of the excessive amount of the 
excretion. This fact is so characteristic that a tentative diag- 
nosis may be made upon this alone. 

From the foregoing it is evident that, from a clinical stand- 
point, it is necessary to consider the specific gravity, the total 
solids, and the total volume together, as they are intimately and 
logically related to each other, and that the determination of 
the specific gravity is of greatest value and significance when 
the total volume of the urine for twenty-four hours is known. 
Also that the chief clinical value of the study of the amount and 
of the specific gravity is to aid in the estimation of the total 
solids of .the urine (see below). 

The specific gravity of the urine is usually determined with 



SPECIFIC CKAVITY. 



169 



the aid of a. hydrometer or urinometer, but is more accurately 
determined by the Westphal balance. Only approximately cor- 
rect results may be obtained with the urinometer. 

The Use of the Urinometer. — The scale of the urinometer 
is usually marked in regular intervals from 1000 to 1060 (Fig. 
20). To insure ease and accuracy in reading, these markings 
should not be too close together. Many urinometers are 
inaccurately made, so that before purchasing an instrument it 
is always well to compare it with a standard instrument, or at 
least to ascertain that it floats at the 1000 mark in distilled water 
at the standard temperature (15° C. or 60° F.). Although a 




Fig. 20— Various Forms of Urinometers and Urinometer Cylinders. 



large instrument is more accurate, a small one, requiring less 
urine, is more convenient and must frequently be employed 
for lack of sufficient urine in which to float the larger instru- 
ment. 

To determine the specific gravity of the urine : After allow- 
ing the urine to cool to room temperature (about 60° F.), it is 
poured into the urinometer tube (the urinometer and an appro- 
priately sized glass cylinder are usually sold together), and the 
urinometer immersed in the urine ; then with the eye on a level 
with the surface of the urine, the division of the scale is read 
off which corresponds to the lowest part of the curve of the 
meniscus. To insure accuracy the containing cylinder should 
be sufficiently large to allow the urinometer to float freely and 
not come in contact w-ith the sides. All bubbles and froth should 



170 



THE URINE. 



be removed from the surface of the fluid by means of filter-paper. 
If the urine is cooler than 15° C, one-third of a urinometer 
unit should be subtracted for every degree centigrade below the 
standard temperature. If warmer an appropriate addition 
should should be made. Since the specific gravity of individual 
urine specimens vary greatly during twenty-four hours, it is nec- 
essary that the specific gravity should be taken from a mixed 
specimen of the twenty-four hours' collection. A reading taken 
from any single specimen is of little clinical significance. 

If the specimen is too small to work with in the ordinary 
way, it is a very simple matter to dilute with a known propor- 
tion of distilled water. Estimate the specific gravity of the 




Fig. 21.— Westphal Balance. (A. H. Thomas.) 



diluted urine and then calculate the specific gravity of the spe- 
cimen at least approximately. 

The Westphal balance (see Fig. 21) is an extremely ac- 
curate method of determining the specific gravity, carrying the 
reading to the fifth figure (fourth decimal). The method of 
employing the Westphal balance is as follows : — 

When the instrument is mounted the glass plummet which 
is suspended from one end of the beam will balance in distilled 
water at 15° C. and represents 1. Now pour the urine into the 
jar until the twist in the platinum wire is below the surface, 
Weighing is accomplished as follows : Place the first rider upon 
the end of the beam above the float, then place the second rider 
in the first notch to the left on the scale on the beam. If the 
plummet rises place the rider on the second notch. If now the 



METHOD OF ESTIMATING TIN-: TOTAL SOLIDS. 171 

beam balances and the temperature of the urine is 15° C, the 
specific gravity of the urine is exactly 1020. If the float still 
rises take the third rider and find the notch in which the beam 
balances or nearly so. If the beam balances with the third rider 
in the forth notch, the specific gravity is exactly 1024. Should, 
however, the float still slightly rise, take the fourth rider and 
find the exact balance, and if this is in the sixth notch the 
specific gravity is exactly 10246. In other words, the second rider 
(in size) gives the third figure, the third rider the fourth (third 
decimal), and the fourth rider the fifth figure or the fourth 
decimal. With a little practice determinations with the West- 
phal balance are rapidly and accurately made. Care should be 
exercised to see that no air bubbles become attached to the corcl 
or float, and that temperature corrections are made as directed 
below. 

Corrections for Temperature. — The temperature of the 
urine immediately after being voided, ranges from 85° to 95° 
F. (29.5° to 35° C.) ; therefore in taking the specific gravity 
of fresh urine its temperature must be observed, and for every 
seven degrees Fahrenheit that the thermometer indicates above 
the temperature at which the instrument is standardized, one 
degree should be added to the specific gravity, as indicated by 
the instrument. 



METHOD OF ESTIMATING THE TOTAL SOLIDS. 

The accurate chemical method of weighing is too difficult 
and tedious for general clinical work, descriptions of which may 
be found in works on physiologic chemistry. 

The normal average weight of solids is sixty-five grams. 
The specific gravity is a pretty accurate index of the amount of 
solids excreted by the urine w T hen this has been determined from 
the twenty-four hours' urine. 

The solids excreted in one litre of urine may be approxi- 
mated in grains by multiplying the last two figures of the spe- 
cific gravity by 2.2337 (Vierodfs factor). 

Trapp's Formula. — Multiply the last two figures of the 
specific gravity by two, and the result will represent the parts 
of solids in one litre of urine. Example : If the specific gravity 



172 THE URINE. 

is 1023, then 23 times 2 will equal -±6 parts of solids per 1000 
cubic centimeters. 

Bird's Formula. — The last two figures of the specific gravity 
about represent the grains of solids in a fluidounce of urine 
tested. Thus, a specific gravity of 1022 would contain approxi- 
mately twenty-two grains of solids per ounce of urine. 

Metz's Formula. — Multiply the last two figures of the spe- 
cific gravity by .00233, and multiply this product by the total 
twenty-four hours 5 volume in cubic centimeters. The final 
product will be the total weight of solids expressed in grams. 
Example: Specific gravity = 1024: then 24 x .00233 x 1500 
cubic centimeters equals 87.27 grams of solids in twenty-four 
hours 5 excretion. 



THE REACTION. 

The reaction of the twenty-four hours 5 urine is usually acid, 
sometimes amphoteric, and rarely alkaline. The normal acidity 
is not due to the presence of free acids, but to acid salts, chiefly 
acid phosphates. Primarily the reaction of the urine depends 
upon the character of the diet. Increase in ingestion of food- 
stuffs containing alkaline salts (vegetables), or when such salts 
are formed within the body from organic acids contained in the 
food (fruits), will tend toward an alkaline reaction when the 
urine is voided. The elimination of alkaline urine in this in- 
stance is due to the presence of a fixed alkaline, and therefore 
cannot be considered pathologic. 

It has already been stated that the acid reaction of urine is 
normally due chiefly to the acid phosphates. Besides these a 
certain amount of the neutral phosphates of calcium, ammonium, 
and of sodium usually occur, and it may happen that the acid 
and neutral phosphates are present in such proportions that the 
sample will turn blue litmus red and red litmus blue. Such a 
urine is said to be amphoteric. 

Urine, when allowed to remain exposed to the air at room- 
temperature, gradually undergoes ammoniacal fermentation. 
This is due chiefly to the action of certain micro-organisms upon 
urea, which is decomposed by a hydrolvtic process into ammonia, 
carbon dioxid, and water (see page 162). 



THE REACTION. 173 

The result of this development of an alkaline reaction is 
that the soluble phosphates of the alkaline earths are precipi- 
tated as tri-calciuni phosphate and amnionio-magnesiuni phos- 
phate, at the same time the soluble urates are transformed into 
the insoluble ammonium salt (see Fig. 23). 

Test. — In order to determine whether the alkalinity of a 
given specimen is due to a fixed or to a volatile alkali (am- 
monia ) ? a strip of red litmus is clamped into the cork of the 
Lottie so that it does not touch the liquid but remains dry. 
Volatile alkali will gradually turn the paper blue, while a fixed 
alkali will not affect the paper unless the alkaline fluid touches it. 

In specimens where the degree of akalinity or acidity is so 
slight that the litmus-paper test leaves the examiner in doubt, 
the uncertainty may be eliminated by the following expedient: 
Take two pieces of litmus-paper, one red and the other blue; 
allow the urine to come in contact with but half of each strip 
of paper; then with distilled water allow^ the whole extent of 
both strips to become wet. This eliminates any change in the 
paper due to the presence of moisture alone, and brings out 
sharply slight changes in the color of the litmus-paper by 
allowing comparison between the adjacent portions of each strip 
of paper. 

Increased Acidity. — The urine is frequently found hyper- 
acid in: — 

1. Fevers. 

2. Inflammations of the liver. 

3. Hyperchlorhvclria. 

4. Acute rheumatism. 

5. Lithemia. 

6. Xeurasthenia. 

Alkaline urine may be passed under the following condi- 
tions : — 

1. Undue retention of urine in the bladder. 

2. When there is residual urine. 

3. Presence of urea-decomposing bacteria in the bladder. 

4. Chlorosis. 

5. Organic nervous diseases. 

6. In marked degrees of general debility. 



174 THE URINE. 

ESTIMATION OF ACIDITY OR ALKALINITY OF 
THE URINE. 

Reagents. — (a) A one-tenth normal solution of NaOH 
(1 cubic centimeter equals .00-1 gram NaOH). 

(b) A one-tenth normal solution of HC1 (1 cubic centi- 
meter equals .00365 gram HC1). 

Since these solutions are difficult and tedious to prepare, it 
may be well to obtain them ready-made from reliable chemical- 
supply houses. 

The Test. — If the urine has an acid reaction, measure 100 
cubic centimeters into a beaker, and add a few drops of a litmus 
solution. Now add from a burette, drop by drop, sufficient of 
the NaOH solution to produce an alkaline reaction. State the 
acidity in this wise : 100 cubic centimeters of urine requires 
"F cubic centimeters of one-tenth normal NaOH to render 
alkaline to litmus. Put in place of "X" the number of cubic 
centimeters of NaOH used in the examination. 

If the urine to be tested is alkaline, then first add to the 
100 cubic centimeters exactly 20 cubic centimeters of the one- 
tenth normal HC1 solution. Stir well, and then proceed as in 
case of the acid urine. State the result as follows: 100 cubic 
centimeters of urine requires "Y" centimeters of one-tenth nor- 
mal HC1 to render alkaline to litmus. Put in place of "Y" the 
difference between 20 cubic centimeters and the number of 
cubic centimeters of NaOH used. 

CHEMICAL COMPOSITION OF THE URINE. 

A general idea of the chemical composition of the urine and 
of the quantitative variation of the individual components, may 
be had from the accompanying table (after Simon). The indi- 
viduals from whom the urine was obtained were adults. Their 
habits, diet, etc., may be taken to be that of the average Ameri- 
can city dweller. 

It must be borne in mind that tabes of Chemical composi- 
tion are based upon averages taken from a large number of 
complete analyses, the composition of which, even in perfect 
health, cover a greater range of variation than indicated in the 
following table: 



INORGANIC CONSTITUENTS. 175 



CHEMICAL ANALYSIS OF THE URINE (Simon). 

( trams. 

Water 1200 to 1700 

Solids 60 

Inorganic solids 25.0 to 26.0 

Sulphuric acid (H 2 S0 4 ) 2.0 to 2.5 

Phosphoric acid (PoO-J 2.5 to 3.5 

Chlorine (XaCl) 10.0 to 15.0 

Potassium (K 2 0) 3.3 

Calcium (CaO) 0.2 to 0.4 

Magnesium (MgO) 0.5 

Ammonia ( NH 3 ) 0.7 

Fluorids, nitrates, etc 0.2 

Organic solids 20.0 to 35.0 

Urea 20.0 to 30.0 

Uric acid 0.2 to 1.0 

Xanthin bases 1.0 

Creatinin 0.05 to 0.06 

Oxalic acid 0.05 

Conjugate sulphates 0.12 to 0.25 

Hippuric acid 0.65 to 0.7 

Volatile fatty acids 0.05 

Other organic solids 2.5 



THE INORGANIC CONSTITUENTS OF 
THE URINE. 

The inorganic constituents of the nrine represent the excess 
of mineral salts that find their way into the blood from the 
digestive tract or which develop within the body during the 
processes of metabolism, especially during albumin decomposi- 
tion. We therefore find that exercise and the ingestion of large 
amounts of food, as well as the increased cell activity occurring 
in acute fevers, lead to an increased elimination of salts, and 
conversely smaller amounts of salts are eliminated when the 
intake of food in general is restricted. These statements apply 
particularly to the phosphates. 

The bases which are found in the urine in combination with 
hydrochloric acid, phosphoric acid, and sulphuric acid, are chiefly 
sodium, potassium, calcium, magnesium, and irmmonium. It is 
believed that the mono-acid phosphates of the alkaline earths 
are held in solution by sodium chlorid, and also by the di-acid 
sodium phosphates, to which latter the acidity of the urine is 
largely due. 

While the greater portion of the sulphuric which results 



176 THE URINE. 

from albumin decomposition is found in the urine combined with 
inorganic bases, a variable fraction also occurs united with cer- 
tain aromatic substances which are developed in the intestines 
during putrefaction and decomposition. The resulting bodies 
are spoken of as the ethereal or conjugate sulphates. They com- 
prise the alkaline salts of indole skatol, and phenol. 

The Phosphates. — The amount of phosphoric acid excreted 
by the healthy individual in twenty-four hours ranges from 2.3 
to 3.5 grams, the average being 2.8. In the urine the phosphoric 
acid is found in part combined with the alkaline earths — earthy 
phosphates — and in part with the alkalies as the alkaline phos- 
phates. The alkaline bases represent about two-thirds of the 
total phosphoric acid in combination. The earthy phosphates are 
insoluble in water, but are soluble in dilute acids. They comprise 
the phosphates of calcium and magnesium. The calcium phos- 
phates predominate. In acid urine the earthy phosphates are in 
solution, while in alkaline urine they are precipitated. They are 
thrown down by heat, and also when the reaction of the urine 
becomes alkaline, whether from a course of internal alkaline 
medication or from the putrefaction of the urea of the urine. 
If, during decomposition of the urine, the contained acid phos- 
phates are acted upon by the ammonia resulting from urea 
decomposition, ammonium-magnesium phosphate (triple phos- 
phate) is formed and appears in the urine as the characteristic 
prismatic or coffin-lid crystals. 

The alkaline phosphates comprise the phosphates of sodium 
and potassium; of these the sodium is the more abundant. 
These, unlike the earthy phosphates, are easily soluble in water 
and in alkaline fluids. The alkaline phosphates form the 
chief bulk of urinary phosphates. The normal urinary acidity 
depends upon the presence of these acid phosphates, and not 
upon the presence of free acid. 

While the bulk of the phosphates in the urine is derived 
from the decomposition of food-stuffs, a part also is derived 
from the breaking down of the highly complex organic bodies: 
lecithin and nuclein. 

Clinically, the execretion of phosphoric acid and its determi- 
nation is of very little significance, since it is so largely dependent 
upon the influence of diet, exercise, etc. 



INORGANIC CONSTITUENTS. 177 

Detection. — If neutral or alkaline urine is heated in a 
test-tube, a precipitate will be formed which will be found to 
consist of earthy phosphates. Such a precipitate might be caused 
by the presence of albumin. To remove the doubt add a few 
drops of dilute (10 per cent.) acetic acid, when a precipitate due 
the phosphates will immediately disappear, while if albuminous 
the cloud will remain or may be increased. 

Test for Earthy Phosphates. — To a few cubic centi- 
meters of urine in a test-tube add a few drops of liquor potassii 
and boil. The earthy phosphates will be thrown out of solu- 
tion, and may, after settling, be collected upon a filter. Now, 
to the filtrate add one-third of its volume of "magnesia mix- 
ture" (for formula see Appendix). The precipitate thus formed 
represents the phosphoric acid that was in combination with 
the alkaline bases, combined now^ in the form of ammonium- 
magnesium phosphate. 

Estimation or the Phosphates by the Centrifuge. — 
In a graduated percentage centrifuge tube (see Fig. 30) mix 10 
cubic centimeters of urine with 5 cubic centimeters of "magnesia 
mixture"; invert several times to thoroughly mingle. Kevolve 
in the centrifuge (Figs. 26, 27 and 28) for three minutes. Head 
off every one-tenth cubic centimeter of precipitate as 1 per cent, 
by bulk of total phosphates. The average normal percentage by ' 
this method is eight. Eoughly, each one-tenth cubic centimeter 
of sediment is equal to about 0.0225 per cent, bv weight of 
P 2 5 . 

Determination of the Total Phosphoric Acid. — For 
this determination the following solutions are required : — 

1. A standard solution of uranium nitrate: 20.3 grams of 
uranium nitrate are dissolved in 1000 cubic centimeters of dis- 
tilled water, then each cubic centimeter of this mixture is 
equivalent to 5 milligrams of phosphoric acid. 

2. Sodium acetate solution: 100 grams of sodium acetate 
are dissolved in 900 cubic centimeters of distilled water, and to 
this 100 cubic centimeters of acetic acid are added. 

3. Saturated solution of potassium ferrocyanide. 
Method. — Fifty cubic centimeters of urine are placed in a 

beaker and 5 cubic centimeters of the sodium acetate solution 
added. The mixture is warmed over a water-bath and uranium 

12 



178 THE URINE. 

acetate solution added from a burette as long as a precipitate 
is formed. If the formation of precipitate is not easily recog- 
nized, a drop of the potassium ferrocyanide solution may be 
added; then, as long as a brown color does not appear where 
the drop falls, the uranium nitrate solution should continue to 
be added. The end point in the precipitation reaction is 
reached when a reddish-brown discoloration appears upon the 
addition of a drop of the uranium nitrate. The quantity of 
uranium nitrate solution employed to accomplish complete pre- 
cipitation is now read off from the scale on the burette, each 
cubic centimeter of which will equal 5 milligrams of phosphoric 
acid. This number and the 50 cubic centimeters of urine em- 
ployed furnished the working bases for calculating the per- 
centage. 

The presence of sugar or albumin does not interfere with 
this reaction. 

Separate Estimation of the Earthy and Aleaxine 
Phosphates. — Two hundred cubic centimeters of urine in a 
beaker are rendered alkaline by the addition of ammonium 
hydroxid, and set aside for a few hours. The earthy phos- 
phates are thus precipitated and may be collected upon a filter- 
paper, and after washing with dilute ammonia (1 : 3) are trans- 
ferred to a beaker, where they are dissolved with as little acetic 
acid as possible. Distilled water is then added so as to make 
the total volume approximately 50 cubic centimeters, when the 
solution is boiled and then titrated as above. In a second por- 
tion of urine the total phosphates are determined as outlined 
above. Then the difference between the two results will repre- 
sent the quantity of phosphoric acid present in combination 
with the alkalies. 

Significance. — As has been shown above, the phosphates are 
dependent upon many uncertain factors which are determined 
with difficulty and are of little clinical significance. When 
in a given case the phosphates are constantly thrown out of 
solution, it may be taken as an indication that the formation of 
gravel or calculus is impending. If the usual signs of ammo- 
niacal fermentation are present, the significance is plain. 



THE SULPHATES. 179 

THE SULPHATES. 

General Considerations. — The major portion of the sul- 
phates appearing in the urine are derived from the food, and 
comprise the simple mineral sulphates of sodium and potassium. 
Only a small portion exists in organic combination as the ethe- 
real or conjugate sulphates. 

The three predominating conjugate sulphates are: Phenol 
potassium sulphate, indoxyl potassium sulphate (indican), and 
skatoxyl potassium sulphate. 

The mineral sulphates comprise about nine-tenths of the 
total sulphates in the urine. 

Test for Mineral or Preformed Sulphates. — Add a few 
drops of acetic acid (to prevent the formation of barium phos- 
phate) to a test-tube half full of urine. Now, upon the addition 
of a solution of barium chlorid a white precipitate of insoluble 
barium sulphate will be formed. This precipitate varies in 
density from a faint white cloud to a bulk that gives a thick 
creamy consistence to the whole mixture. One can roughly deter- 
mine by the amount of precipitate as compared with the known 
normal standard, whether the sulphates are increased or not. 

Test for Conjugate or Ethereal Sulphates. — Mix 
equal quantities of alkaline barium chlorid (see Appendix) and 
urine in a test-tube. After allowing a few minutes for the 
precipitate to form the mixture is filtered. This process precipi- 
tates both phosphates and preformed sulphates. The filtrate is 
now acidified with 5 cubic centimeters of a (one-fifth vol.) HC1 
solution and then boiled for some time. The presence of ethe- 
real sulphates is indicated by a reddish discoloration of the 
fluid which in addition becomes turbid. 

Potassium Indoxyl Sulphate or Indican. — As this substance 
represents the characteristic ethereal sulphate, it may be taken 
as an indicator for the w T hole group, and the tests for this sub- 
stance used for estimating the relative amount of the whole 
group of ethereal sulphates in the urine. The several tests for 
indican are based upon the fact that an excess of HC1 will 
liberate the indoxyl, which can then, by the addition of an 
oxidizing agent, be converted into indigo-blue, and finally this 
can be recognized in small amounts by extraction from the bulk 
of urine with chloroform. 



180 THE URINE. 

Test for Indican (modified Jaffee). — Take 20 cubic centi- 
meters of filtered urine in a test-tube and add 3 or 4 »cubic 
centimeters of chloroform, and one drop of a 1 per cent, solu- 
tion of potassium chlorate, and finally 20 cubic centimeters of 
HC1. This mixture is to be thoroughly agitated and mixed by 
pouring repeatedly from one test-tube to another. This should 
be repeated at intervals of two or three minutes covering a 
period of ten minutes. The presence of indican is indicated by 
a blue discoloration of the chloroform. In the presence of a 
large amount of indican the chloroform will appear almost 
black, while the whole mixture will assume a dusky bluish-red 
color. (For comparative color-scale see Plate III.) 

While a trace of indican cannot be considered pathologic, 
there are, nevertheless, many specimens, possibly one-third, 
which fail to show any discoloration of the chloroform. 

Caution. — An excess of oxidizing agent, either in volume 
or strength, will prevent a positive reaction through over- 
oxidation of the indoxyl compound. 2. An excess of chlo- 
roform will result in too great dilution of the indigo, causing 
failure to detect traces of indican. 3. A reddish discoloration 
of the chloroform is not due to indican. Such a reaction occurs 
in the urine of patients who are taking iodide, and possible bro- 
mide. The simultaneous occurrence of a red and a blue re- 
action, will produce a color bordering on purple. This possible 
source of error may be removed by the addition of a few drops 
of a 10-per-cent. solution -of sodium thiosulphite, which will 
bleach to pink color clue to iodine. 

Clinical Significance of Indican. — The presence of 
more than a trace of indican probably denotes the existence of 
undesirable decomposition in the intestinal tract. The occur- 
rence of a positive indican reaction does not necessarily indicate 
constipation. 

THE CHLORIDS. 

The quantity of chlorids in the urine is usually decreased 
in: 1. Most febrile diseases. 2. Xephritis. 3. Wasting dis- 
eases. 4. Pneumonia. 

Approximate Estimation of the Chlorids. — To a few cubic 
centimeters of urine in a test-tube, add a few drops of nitric 



* o 5 
>j2 














W 2 ^ 



Q ^ 



« o 



ORGANIC CONSTITUENTS. 181 

acid, boil and filter to remove the albumin. Add to the filtrate 
a Tew drops of a 10-per-cent. solution of silver nitrate. The 
abundance of the white cloud will roughly indicate the amount 
of chlorids present. 

Accurate Methods of Quantitative Determination. — Method 
of Salkowski-Yolhard. Take 10 cubic centimeters of urine in 
a beaker and dilute with 50 cubic centimeters of distilled water, 
and then treat with -i cubic centimeters of concentrated nitric 
acid and 15 cubic centimeters of a standard silver nitrate solu- 
tion. The mixture is then further diluted with distilled water 
up to 100 cubic centimeters, and after thorough agitation is 
passed through a dry filter. In a carefully measured portion of 
the filtrate the excess of silver is carefully titrated with a solu- 
tion of potassium sulphocyanide of such a strength that 25 
cubic centimeters corresponds to 10 cubic centimeters of the 
standard silver solution. A few drops of a saturated solution 
of ammonio-ferric alum serve as an indicator. The amount of 
silver solution employed to precipitate the chlorids in the 10 
cubic centimeters of urine is then calculated. The number of 
cubic centimeters necessary for this precipitation is multiplied 
by 0.01, which will indicate the amount of chlorids in 10 cubic- 
centimeters of the urine calculated as sodium chlorid. The 
presence of albumins or of sugar does not interfere with this 
reaction. 



THE ORGANIC CONSTITUENTS OF THE URINE. 

GENERAL CONSIDERATIONS. 

The organic constituents of the urine comprise the normal 
end-products of nitrogenous metabolism wdthin the body, also 
various products of albuminous putrefaction which have found 
their way into the general circulation from the intestinal tract. 
Finally certain pigments which bear a relation to the normal 
blood-pigment, and various substances of obscure origin are en- 
countered. Under abnormal conditions we may meet with 
normal constituents of the blood wmich do not ordinarily appear 
in the urine. Lastly, in pathologic conditions, we meet various 
products of abnormal metabolism. 



182 THE URINE. 

UREA. 

Urea was first synthetically prepared from ammonium cya- 
nate in 1828 by Wohler. Formerly it was supposed that urea 
resulted from uric acid through a process of oxidation, and that 
this was its only source. Now this error is well known, and 
while it is recognized that the formation of urea from uric acid 
is possible and that a small portion of the total amount may be 
derived in this manner, modern research has shown that in man 
urea is largely derived from the destruction of the nucleins 
within the body, and that the sources of the nucleins are both 
the tissue cells and the cells of animal foods ingested. 

It has been repeatedly shown that during the decomposi- 
tion of albumins by means of acids and alkalies, as during the 
process of tryptic digestion and albuminous decomposition, a 
large amount of mono-amido-acids results. And it is supposed 
that these bodies probably represent the intermediary products 
in the transformation of albuminous nitrogen into urea. Under 
certain pathologic conditions these acids may appear in the 
urine, and when they are there noted the elimination of urea is 
much diminished. In health, however, this does not occur, and 
on examination of the different tissues of the body such acids 
are found only in traces. We must conclude, therefore, that 
these acids, supposing them to occur as the primary products 
of albuminous decomposition within the body, are transformed 
at once into other substances, which in turn give rise to urea. 

It has also been shown that the amido acids yield carbamic 
acid on oxidation, and that on alternate oxidation and reduction 
urea can be produced from the ammonium salt (ammonium 
carbamide). While it is generally assumed that urea is largely 
referable to a transformation of the mono-amido-acids into 
ammonium carbamate, and while it has also been shown that 
such a transformation does actually occur, it must be remem- 
bered that at best only traces of these amido acids are found in 
the tissues. 

From these conclusions it is reasonable to believe that the 
greater portion of albuminous nitrogen is set free from the vari- 
ous organs of the body in the form of an ammonium salt of para- 
lactic acid, and that this salt is now generally conceded to be 
the antecedent of urea. 



UREA. 188 

It is probable that a certain amount of urea is produced 
in the body in a number of ways, and there is ground for tin- 
belief that its formation is not confined to a single organ. 
The greater part is, without doubt, produced in the liver. In 
corroboration of this fact it has been repeatedly shown that in 
disease of this organ, when accompanied by extensive destruc- 
tion of the glandular elements, a diminished amount of urea is 
found in the urine, while ammonia and lactic acid are found 
in increased amounts. In certain cases of this class as much 
as 37 per cent, of the total urinary nitrogen has been found in 
the urine in the form of ammonia. 

If we accept the modern doctrine that urea originates not 
only in different ways, but that it may also be formed in other 
organs beside the liver, then we can understand why it is that 
in certain diseases of the liver the diminution in the amount of 
urea excretion is not always proportionate to the extent of 
parenchymatous degeneration, and that no case has yet been 
reported in which the excretion of urea has ceased altogether. 

Nitrogenous Equilibrium. — The albumins are the ultimate 
source of the urea, and according to Pettenkof er they exist within 
the body in two forms, viz. : as organized albumin, which is built 
up into tissues, and the so-called circulating albumin, which is 
present in excess of the actual requirements of the body and 
which may be broken down directly and eliminated in the urine 
without ever having entered into the construction of the body- 
proper. This latter portion of the body albumin is said to fur- 
nish the bulk of the urea, while the fixed tissue albumin repre- 
sents the minor but more uniform source. The actual amount 
eliminated will, therefore, be primarily dependent upon the quan- 
tity ingested. 

Experiment has shown that under normal conditions of 
average diet the total urinary nitrogen is practically equivalent 
to the quantity ingested, barring a small fraction, which escapes 
digestion and appears in the feces. Such a condition is spoken 
of as the nitrogenous equilibrium of the body. Of this relation 
infinite variations exist, which may even vary from time to time 
in the same individual. If the amount of nitrogenous food is 
suddenly diminished the amount of urinary nitrogen will also 
decrease; then if the reduced ingestion remains constant the 



184 THE URINE. 

nitrogen output, while lowered, will at the same time tend to 
remain level. If, on the other hand, the nitrogen intake is in- 
creased, an increased nitrogen elimination speedily follows, 
but here a certain fraction will be retained in the body and 
gradually a higher level of equilibrium will be established. 

There are natural limits to this power of accommodation 
of the system to nitrogenous ingestion and elimination, so that 
we may find a point which varies in different individuals where 
a further nitrogen ingestion does not lead to a higher level of 
nitrogenous equilibrium, and when, consequently, a further re- 
tention of nitrogen does not occur. Overfeeding then results in 
various digestive disturbances. Diarrhea and vomiting may 
occur through nature's effort to protect the body from a further 
increase in circulating nitrogen. 

Underfeeding, on the other hand, generally leads to in- 
creased destruction of the organized albumins. Although for a 
while the body's store of fats and carbohydrates is capable of 
protecting the body against an undue loss of nitrogen in this 
direction, still if the reduced intake of nitrogen continues suf- 
ficiently long, death finally ensues. 

From the fact that the level of nitrogenous equilibrium 
varies in different individuals and in the same individual from 
time to time, it follows that the amount of urea excreted must 
also vary according to the same irregular manner. Any figures, 
therefore, which are supposed to indicate the amount of urea 
eliminated, can be of little and uncertain value unless the actual 
state of the individual's health is known, also his body-weight, 
habits as to exercise, the amount of nitrogenous food ingested, 
etc. Only when we are in possession of an accurate knowledge 
of these several factors can we say whether the urea excretion is 
or is not normal. 

Certain figures have been compiled by physiologists to indi- 
cate the amount of nitrogen which should enter into the com- 
position of the diet and from which we may approximately 
calculate the amount of urea that should be excreted. By esti- 
mating this, or still better, by determining the total nitrogen 
elimination, we can then determine whether or not the indi- 
vidual is consuming the proper amount of nitrogenous food in 
his dietary. While such figures may serve as a general guide, 



UREA. 185 

they have mostly been constructed without due regard for the 
factors above indicated, and should not, therefore, be too im- 
plicitly relied upon. 

According to Simon, among the well-to-do classes the elimi- 
nation of from twenty to twenty-five grams of nitrogen in 
twenty-four hours is about normal, taking the average body- 
weight of the individual into consideration. A smaller amount 
is not infrequently met in persons of sedentary habits who may 
appear to be in perfect health. 

Urea in Disease. — While extensive variations occur in the 
urea excretion of health, still greater variations from the average 
standard are noted in disease. But here also should be taken 
into account the amount of nitrogen ingested in relation to the 
body-weight. 

An increased elimination of urea referable to the destruc- 
tion of organized albumins is frequently observed, but this 
may in cases be obscured by a deficient nitrogen ingestion un- 
less the total intake of the latter is not definitely known. It 
is of interest here to note that in certain diseases of the liver 
in which there is great destruction in the parenchyma, the 
amount of urea may be markedly diminished, even when a fairly 
abundant supply of nitrogen is taken in. This w T ill be found 
due largely to the interference in urea synthesis, and as a sec- 
ondary result we find that in these cases a considerable portion 
of the nitrogen appears in the urine in the form of ammonium 
salts of paralactic acid (see page 182) and of carbamic acid; in 
extreme cases as mono-amido-acids and as leucin and tyrosin. 

Properties of TJrea. — Urea crystallizes in colorless, quadri- 
lateral, or six-sided prisms with oblique ends, or when rapidly 
crystallized in delicate white needles which melt at 132° C, but 
which are probably decomposed at a temperature of 100° C. 
They contain no w r ater of crystallization, and are permanent in 
the air, and easily soluble in cold w^ater, in which they form a 
neutral solution. With nitric acid, urea unites to form urea 
nitrate, w r hich crystallizes out in octahedral lozenge-shaped or 
hexagonal platelets. Urea nitrate is readily soluble in distilled 
water, but is soluble less readily in water acidified with nitric 
acid. Its formation is frequently observed when urine is exam- 
ined cold, with nitric acid, for the presence of albumin. On 



186 THE URINE. 

heating, the crystals are decomposed without leaving any 
residue. 

Detection of Urea. — 1. To detect urea place a drop or two 
of the suspected fluid upon a glass slide, and after adding a drop 
or two of nitric acid, warm gently. If urea is present, after 
partial evaporation and cooling, the microscope will show the 
characteristic crystals of urea nitrate. These are either rhombic 
or hexagonal plates, frequently overlapping like shingles on a 
roof. Their acute angles measure 82°. 

2. Add to the suspected fluid, in a test-tube, a few drops 
of fresh sodium hypobromite. A rapid evolution of gas will 
indicate the presence of urea. 

Since clinical observations are concerned in the total out- 
put of urea, it becomes necessary to estimate the quantity or 
percentage from a sample of twenty-four hours 7 urine. 

Under normal conditions of average health the percentage 
of urea is two. 

The average daily excretion of urea is forty grams or five 
hundred grains, or about half the weight of the total solids. 

Quantitative Estimation of Urea. — This quantitative esti- 
mation is determined by calculation from the observed volume 
of nitrogen gas evolved from a measured quantity of urine by 
a process of decomposition. For practical purposes one gram 
of urea is estimated to furnish 37 cubic centimeters of nitrogen 
gas. The decomposition of the urea contained in the meas- 
ured volume of urine is accomplished by means of an alkaline 
solution of sodium hypobromite (see Appendix for formula of 
Knopfs solution). The test is best conducted in a Doremus- 
Hinds ureameter (see Fig. 22). 

The Test. — First pass some urine through the small tube 
to wet the stop-cock, then fill the large tube with Knop's solu- 
tion, and the smaller side tube with urine exactly up to the 
cubic centimeter mark. Now, drop by drop, allow exactly 1 
cubic centimeter of urine to pass into the reagent in the large 
tube. When the bubbles of gas (nitrogen) cease to rise, the 
fraction of a gram of urea may be read directly from the grad- 
uated scale on the larger tube. 

Example. — Suppose that after admitting exactly 1 cubic 
centimeter of urine the level of the fluid stands at the 0.018 



URIC ACID. 



187 



mark. Then the 1 cubic centimeter of the sample contained 

0.018 gram of urea. To obtain the total amount of urea ex- 
creted in twenty-four hours, it is only necessary to multiply 
this figure by the number of cubic centimeters in twenty-i'our 
hours' urine. 

URIC ACID. 

General Considerations. — Uric acid, like urea, is nitrogen- 
ous. The normal proportion of uric acid to urea is as 1 : 45. In 
health it exists in solution as sodium and as potassium urate. A 
healthy adult excretes 0.2 to 1.0 gram of uric acid in the course 
of twenty-four hours. The amount increases physiologically 




Fig. 22.— Various Forms of Ureameters. 



with increased ingestion of food, and pathologically with in- 
creased nitrogen metabolism in about the same proportions as 
urea. The amount of uric acid in urine varies directly with the 
specific gravity, so that the last two figures of the specific gravity 
(calculated to four places), multiplied by two, give approxi- 
mately the number of centigrams of uric acid in the litre. The 
daily excretion of uric acid is increased in fevers and in leu- 
kemia. The relation of the elimination of uric acid to attacks 
of gout and the so-called uric acid diathesis, is still an unsettled 
question. Of necessity the elimination of uric acid is increased 
by the ingestion of uric acid and other purin bodies, as well as 
by foods rich in nuclein (rich in cells). 

Properties of Uric Acid. — Uric acid is practically insoluble 
in cold water, requiring 18,000 parts of water to dissolve 1 part 



188 THE URINE. 

of uric acid. It is freely soluble in alkalies and in solutions of 
the carbonates. Pure uric acid crystals are colorless, transparent 
platelets, the angles of which are frequently irregular and 
rounded off. Such crystals are occasionally seen in freshly 
voided urines, but more commonly they are found in the 
form of brownish-yellow, whetstone-shaped crystals, occurring 
singly or in groups (see Plate IV, a and b). Uric acid once 
deposited remains undissolved in acid urine. Owing to the 
inclusion of the urinary pigments in the crystals, these, when 
present in appreciable quantity, are spoken of as "brick-dust." 

Microscopic Appearance. — The reddish specks observed by 
the naked eye are found, upon microscopic examination, to 
be various modifications of rhombic prisms (see Plate IV, c, d, 
e and /) . The simpler forms have some resemblance to the con- 
ventional lozenge or whetstone. 

Significance. — If these crystals deposit in appreciable quan- 
tities soon after micturition, it may be considered as a sign of 
impending gout or gravel formation. Such a deposit, however, 
is not necessarily evidence that the elimination of uric acid is 
excessive. 

Isolation of Uric Acid. — To 200 cubic centimeters of urine 
add 10 cubic centimeters of HC1, and let stand for twenty-four 
hours; the uric acid will then have settled to the bottom of 
the container, from which it may be collected by decanting, 
filtering, and finally washing in cold water. 

Qualitative Tests for Uric Acid. — Murexid Test : Put the 
solution supposed to contain uric acid or urates in a porcelain 
dish, add a drop of nitric acid and evaporate to dryness. 
After cooling, allow a drop or two of ammonia water to 
come in contact with the residue. The presence of uric acid 
or urates will be shown by bright blue or violet (murexid) color. 

Schiff's Test. — Having a residue prepared as above, or 
crystals supposed to be uric acid, dissolve in a test-tube with 
the aid of a solution of sodium carbonate. Moisten some 
filter-paper with a 10-per-cent. solution of silver nitrate. Into 
the center of this paper allow a few drops of the uric acid 
sodium carbonate solution to fall. In the presence of uric acid 
the silver nitrate will be reduced to black metallic silver. 

Approximate Quantitative Determination. — According to 



THE URATES. 189 

Gubler 2 the amount of uric acid in urine may be approximately 
determined by stratifying the specimen of urine to be tested 
upon a layer of nitric acid contained in a test-tube, so that the 
volume of urine to the volume of nitric acid is as 3 : 2. After 
a short interval uric acid crystals will separate out as a cloudy 
white ring at the line of junction of the two fluids. If the 
amount of uric acid in the specimen is increased, the precipita- 
tion will be plain in five minutes or less. If diminished it will 
not appear until later. This determination is of value only 
when the daily excretion of urine is approximately normal, and 
if it is diminished in amount it should be diluted with water 
up to the average amount before applying the test. Obviously 
the conclusions derived from such a method should be. given 
very slight weight clinically, and then only when the estima- 
tion is made with a part of the twenty-four hours' collection, 
properly diluted as mentioned above. Albumin must first be 
removed by slight acidulation with dilute acetic acid, boiling 
and filtering, after which the test should be applied when the 
urine has cooled to room-temperature. 



THE PURIN BASES IN THE URINE. 

These comprise xanthin, hypoxanthin, heteroxanthin, para- 
xanthin, guanin, and adenin. These bases are normally present 
in the urine and comprise approximately one-tenth as much as 
the normal uric acid. The amount of purin bases normally 
found in the urine in twenty-four hours varies between 0.028 
and 0.058 gram. There is at present no practical clinical 
method of estimating them. 



THE URATES. 

General Considerations. — When the urine cools the urates 
may settle to the bottom of the container as a cloudy reddish- 
yellow precipitate. This is most likely to occur when the urine 
is scanty, concentrated, and highly acid. This condition often 
obtains in fevers, and in congestion or inflammation of the kid- 



2 Laquer : Schmidt's Jarhbucher. Vol. ccxxxvi, No. 10. 



190 THE URINE. 

neys. The sediment then presents a fairly characteristic appear- 
ance, being clay-colored, reddish-yellow or rose-red. It may 
adhere to the walls of the vessel as a fine, reddish coating. 

The ordinary nratic sediment consists of a mixture of the 
urates of sodium, potassium, calcium, magnesium, and ammo- 
nium. Sodium urate predominates. With the exception of 
ammonium urate, these urates only appear in acid urine. 
Uratic sediments often contain a few crystals of uric acid 
which have been formed during the double rearrangement of 
molecules which resulted in the precipitation of the urates. If 




m 






Fig. 23.— Urinary Sediment. 
a. Triple Phosphates, b. Ammonium Urate in Alkaline Urine. 

urine containing a uratic sediment is decomposed by ammo- 
niacal fermentation, the sediment will be changed to acid-am- 
monium urate, and this latter is the only urate sediment which 
occurs in alkaline urine. 

If freshly voided urine is kept in a cool place, the precipi- 
tation of urates will occur rapidly. This same precipitation 
occurs at a higher temperature if the amount of urates is in 
excess of normal or if the urinary acidity is decreased. This 
precipitate is readily distinguished from phosphates by its 
prompt disappearance upon the application of heat. 

Qualitative Tests. — 1. Half fill a test-tube with turbid urine 
and apply heat to the upper part. If the turbidity is due to the 



CREATININ. 191 

presence of urates, the heated portion of the urine immdiately 
becomes clear. 

2. To some urine in a test-tube add some liquor potassii, 
when the turbidity due to urates promptly disappears. 

Microscopic Appearance of Urates. — The uratic deposit is 
composed of fine, somewhat regular granules, usually occurring 
in groups; sometimes the granules show spiny projections. 
Ammonium urate occurs only in alkaline urine, and is generally 
accompanied by a copious precipitation of triple phosphates. 
Ammonium urate appears as opaque brownish-red spherules with 
or without projecting spines. (See Fig. 23.) 

Significance. — The excess of urates is of no special im- 
portance ; they are increased in most conditions, accompanied by 
fever, and in many disturbances of metabolism. 

HIPPURIC ACID. 

Hippuric acid, in combination with alkaline bases, is a nor- 
mal constituent of the urine. The average quantity eliminated 
in twenty-four hours is one gram. This amount may be in- 
creased by exercise, by a vegetable diet, and by ingestion of ben- 
zoic acid. In cases where the total excretion of urine is greatly 
diminished, hippuric will be spontaneously thrown out of solu- 
tion. This is, however, a rare occurrence, because hippuric acid 
is readily soluble in water. 

Microscopic Appearance. — The crystals are characteristic 
rhombic prisms, resembling in a measure the coffin-lid crystals 
of triple phosphates. Hippuric acid crystals may, however, be 
distinguished by the fact that they are precipitated in acid 
urine only, and also because they do not dissolve on the addi- 
tion of acetic acid. Phosphates, on the other hand, are precipi- 
tated only in neutral or alkaline urine, and are readily soluble 
in dilute acetic acid. 

CREATININ. 

This normal urinary constituent is present in the twenty- 
four hours' specimen to the amount of one gram. Creatinin is 
derived from the creatin of muscle. It is distinguished in the 
urine by its union with mercuric chlorid, with which salt it 



192 THE URINE. 

forms insoluble compounds. Similar characteristic compounds 
are formed with zinc chlorid and silver nitrate. The zinc chlo- 
rid combination has a characteristic appearance, by which this 
substance may be recognized. Under the microscope the zinc 
chlorid combination of creatinin appears as minute needles 
arranged in balls and rosettes. Creatinin reduces alkaline copper 
solutions, and therefore affects, in a slight degree, the accuracy 
of the quantitative sugar estimations which depend upon the 
reducing power of sugar-containing urine. 

OXALIC ACID. 

Oxalic acid is normally present in the urine only in very 
small amounts and it appears in combination with calcium as cal- 
cium oxalate. The normal amount of this substance in a urine of 
normal volume is held in solution by the acid sodium phosphate. 
Since oxalic acid requires 500,000 parts of water to effect solu- 
tion of 1 part calcium oxalate, even the slightest variation in 
the normal relation results in its appearance as a deposit. Arti- 
ficially the crystals may be thrown down by carefully neutral- 
izing the specimen with dilute ammonia water, or by simply 
allowing the urine to stand for a time exposed to the air. The 
sediment of calcium oxalate is usually so scant that it is only 
recognized by the aid of the microscope. The more rapid the 
formation of these crystals in the urine after voiding, the smaller 
will the individual crystals appear; this will, in a way, indicate 
whether or not the presence of the crystals is due to a pathologic- 
increase in the oxalic acid content, or merely to a change 
of reaction (loss of acidity) from standing. The typical cal- 
cium oxalate crystal is a perfect octahedron without color, in- 
soluble in acetic acid, but freely so in hydrochloric acid. (Plate 
V, a and 5.) 

The presence of crystals of the oxalates in the urine does 
not necessarily imply a pathologic increase in the total quantity 
of oxalic acid, since an increase in oxalic acid elimination can 
only be determined by a careful quantitative estimation from the 
twenty-four hours' specimen. It is probably that "oxaluria" is 
often diagnosed on insufficient grounds: a too hasty assump- 
tion, based on the finding of a few crystals, should be guarded 
against. 



LEUCIN AND TYROSIN. 193 

Microscopic Appearance. — Oxalates are recognized in one 
of two forms: perfect octahedra, or in hour-glass and dumb- 
bell forms. 

Significance. — These deposits sometimes follow the eating 
of stewed rhubarb or other acid fruits containing oxalates. 
Usually their existence in freshly-voided urine, indicates imper- 
fect oxidation of retarded metabolism within the economy. 

CYSTIN. 

This substance, in exceedingly small amounts, is a nor- 
mal constituent of urine. It is almost insoluble in water, 
and in the very rare cases where it is present in increased 
amount, it is deposited. To the naked eye the deposit is abun- 
dant and somewhat resembles that of urates. It is, however, not 
dissolved by heat or by the vegetable acids, but is readily dis- 
solved in ammonia water. The ammoniacal solution exposed 
on a slide and examined under the microscope, slowly develops 
crystals in the form of hexagonal plates. (Plate VI, d.) Iodo- 
form, which has found its way into the urine from surgical 
dressings, may be mistaken for cystin. 

Tests. — When urine containing cystin undergoes decompo- 
sition, it develops the odor of hydrogen sulphide. When boiled 
with a solution of lead oxid in sodium hydrate, black lead sul- 
phid is formed. 

LEUCIN AND TYROSIN. 

These are present in the urine in cases of acute yellow 
atrophy of the liver, typhoid fever, and phosphorus poisoning. 
(Plate VI, e and f.) 

To examine for these substances, the urine must be concen- 
trated on a water-bath, and the cooled liquid examined under 
the microscope. 

Microscopic Appearance. — Leucin appears as greenish-yel- 
low spheres with concentric markings or radiating spines. Tyro- 
sin as feathery sheaves, which resemble the frayed end of a rope. 



13 



194 THE URINE. 

PART II. 
ABNORMAL CONSTITUENTS OF THE URINE. 

ALBUMIN. 

General Considerations. — Albumin is probably present in 
minute quantities in all urine, since urine always contains a 
variable number of cellular elements derived from the urinary 
tract. Thus, even under normal conditions, by means of suffi- 
ciently delicate tests, we can always demonstrate the presence 
of albumin. This amount is, however, so small that it returns 
a negative result with the tests commonly employed in the clin- 
ical laboratory for the detection of albumin in urine. There- 
fore the presence of demonstrable amounts of albumin, by the 
tests now to be described, must be considered as variations from 
the normal. The "normal" albumin is nucleo-albumin, and its 
clinical significance is slight, even when present in an appre- 
ciable quantity. 

Several other albumins are found in the urine, of which 
serum-albumin and serum-globulin are of the greatest clinical 
importance, and the term albuminuria is understood to indi- 
cate the presence of one or the other, more often of both, in the 
urine. Other albumins which may be found in the urine, be- 
sides those referred to above, are albumose and fibrin. 

The diagnostic and clinical importance of albumosuria have 
not yet been determined. Traces have been found in many of 
the infectious fevers. A large albumosuria may be significant 
of multiple myelomata, since in this affection large amounts are 
frequently observed. It has also been found in osteomalacia 
and in nephritis. 

Fibrin, when present, indicates the direct entrance of 
blood-plasma into the urinary tract. It must be distinguished 
from blood-clots, which come under the head of hematuria. 

Causes of Transient Albuminurias. — Physiologic or Func- 
tional Albuminuria : The presence of easily demonstrable 
amounts of albumin can hardly be considered normal under any 
circumstances. However, the presence of a functional albu- 



ABNORMAL CONSTITUENTS. 195 

minima is recognized by some authorities. An albuminuria 
does, however, occur, in which the organic change in the kidney, 
if there be any, is so slight and evanescent as to be unimportant. 
This form of albuminuria, compared with that caused by severe 
and permanent kidney change, may, with propriety, be termed 
functional, but never, physiologic. 

Functional. — Albuminuria may occur after violent exer- 
cise, after a cold bath, after severe mental strain, and after over- 
eating of proteid food-stuffs, particularly eggs. 

Whether or not the disturbance giving rise to the albumin- 
uria is insignificant and transient (renal hyperemia or anemia), 
or partakes of the gradual and progressive variety due to perma- 
nent alteration in kidney structure, to which the term chronic 
nephritis is applied, must be determined by the history, symp- 
toms, and progress of the case. 

Fevers. — Here the occurrence of albumin in the urine is, 
in all probability, dependent largely upon the intensity of the 
infective process, causing irritation of the kidney epithelium, 
incident to the passage of toxic substances circulating in the 
blood, through the glomeruli and tubules of the kidneys; the 
inflammatory process so induced being indicated by the amount 
and duration of the albuminuria. 

Specific Blood-Changes. — The nature of the changes in 
the blood occurring in scurvy, peliosis, purpura, hemophilia, and 
pernicious anemia, is obscure in relation to the production of 
albuminuria. 

Foreign Substances in the Blood. — This form of albu- 
minuria is seen after the excessive ingestion of lead, mercury, 
ether, chloroform, etc. These probably produce the albuminuria 
by irritation of the kidney through efforts of that organ at 
elimination of the substances themselves, or their oxidation 
products. 

Circulatory Disturbances. — Here the passive congestion 
occurring in cardio-vascular and pulmonary diseases directly 
affects the permeability of the renal epithelium. 

Xeurotic. — This term applies to albuminuria occurring 
during or immediately succeeding attacks of apoplexy, migraine, 
tetanus, delirium tremens, and certain head injuries. It is prob- 
ably toxic in origin. 



196 THE URINE. 

Extra-Renal. — The presence of blood, pus, chyle or 
lymph, which has entered the urinary tract in appreciable quan- 
tities, will cause albumin to appear in the urine. Recently 
attention has been called to a form of albuminuria resulting 
from abnormalities in the function of the genital tract. This 
form comes and goes irregularly, and does not show the pres- 
ence of spermatozoa. 2 

In the majority of the above conditions the quantity of 
albumin is small and varies from day to day. Under these cir- 
cumstances the relative clinical importance of the finding will 
be determined by microscopic examination of the urine, aided 
by the history and course of the disease in which it occurs. 

These small and transient albuminurias are of considerable 
importance from the standpoint of life insurance, since too much 
importance is frequently placed on the finding of a trace of 
albumin, particularly in a single specimen, which is usually 
voided succeeding a full meal. 

Qualitative Tests for Albumin. — In detailing the following 
methods for determining the presence of albumin in the urine, 
no attempt will be made to offer a great variety of tests. The 
principle involved in the majority is the same, and depends on 
the coagulation of albumin in the specimen by heat or by chem- 
ical reagents. 

Before proceeding to make the tests, the urine must be 
filtered or centrifugated to remove the turbidity. If this is due 
to large numbers of bacteria it cannot be entirely removed by 
this process. The persistence of a bacterial turbidity will inter- 
fere with the delicacy of the tests. 

1. Boiling and Acetic Acid. — This is the best and sim- 
plest test for routine laboratory work. If albumin is found it is 
well to corroborate the finding by another method. The urine 
should not be alkaline in reaction. 

The Test. — Fill a narrow test-tube almost to the top with 
clear urine, boil the upper third, being careful to avoid too much 
agitation. In the presence of albumin, or the neutral earthy 
phosphates, or of both, a white cloud will appear in the heated 
area. ISTow, upon the addition from a pipette of a few drops of 



2 William Glenn Young : New York Med. Jour,. Jan. 5. 1907. 



ALBUMIN. 197 

10 per cent, acetic acid, the cloud, it' duo to phosphates, will 
vanish, while it' caused by albumin it will remain and will per- 
haps be increased in density. 'Hie chief advantages of this 
method are its simplicity and delicacy, and the fact that it allows 
a comparison between the treated and untreated urine in the 
same tube. The test is most satisfactory when performed in 
strong daylight with the tube held close to a dull-black back- 
ground. 

Caution. — At times the addition of more acid to a urine 
already acid will convert the albumin into an acid albuminate, 
which is not coagulable by heat. To overcome this the acetic 
acid should always be dilute (10 per cent.), and should never 
be added to acid urine before boiling, and then only in suffi- 
cient quantity to produce the desired reaction. 

2. Xitkic Acid by Contact. — This test is best performed 
with the Meeker albumin tube. 

The Test (with the Meeker tube). — Allow about one-half 
inch of warm, clear urine to enter the tube through the small 
curved tip. Place the forefinger over the upper end of the tube 
to retain the urine, and dry with care the outside of the tube; 
plunge it immediately into nitric acid of sufficient depth 
to allow the acid to flow in and elevate the urine above it. If 
this is done with care the line of demarkation between the urine 
and the acid will be very sharp, and the presence of albumin will 
be shown by a white line at the plane of contact. Five minutes 
should be allowed for the appearance of the reaction, the delicacy 
of which is increased if both the urine and the acid are pre- 
viously warmed to about 60° C. (a temperature too high to be 
endured by the hand for more than a few seconds). 

Sources of Error. — If a cloud appears at the line of con- 
tact in a urine known to contain an excess of urates, this source 
of error can be eliminated by diluting the urine with one or two 
volumes of water, and repeating the test. 

Xitric acid precipitates certain resins which appear in the 
urine after it has been administered for medicinal purposes. 
If these cannot be excluded another test should be employed. 
If an excess of phosphates exist they may be held in solution 
by the addition of a few drops of a 10-per-cent. solution of 
acetic acid. 



198 THE URINE. 

Nucleo-albumin may be held in solution by the addition of 
one-sixth volume of a saturated solution of NaCl, and then pro- 
ceeding as before. 

Purdy's Test. — Mix a half inch of Purdy's reagent (see 
Appendix) with four inches of urine in a six-inch test-tube; if 
a whitish cloud appears either at once or after standing, the 
urine contains albumin. 

Tanret's Test. — Acidify a quantity of urine with dilute 
acetic acid, if mucin produces a cloud, filter and to the filtrate 
add about 5 cubic centimeters of alcohol and heat slightly. 
Now substitute this prepared urine for the urine used in the 
description of the nitric acid contact-test, using Tanret's 
reagent in place of the nitric acid (see Appendix); the pres- 
ence of a white line at the level of contact denotes albumin. 

Quantitative Estimation of Albumin. — Method of Esbach : 
For clinical purposes as a ready means of comparing the albu- 
min content in a given case or a number of cases, this method 
will be found applicable and convenient. 

It must be borne in mind that the resulting amount of 
albumin, which is commonly spoken of as the percentage of 
albumin, means the percentage by bulk of the moist, gravitated 
precipitate, and should never be confounded with the actual per- 
centage of albumin as determined by accurate weighing methods. 
This never amounts to more than 4 or 5 per cent,, while by 
the method of Esbach the moist precipitate may exceed half of 
the bulk of the fluid in the tube. 

The Test. — Fill the albuminometer tube (see Fig. 24) 
with urine to the line marked "U," and then fill with the 
Esbach reagent (see Appendix) to the line marked "K." Thor- 
oughly agitate and then stand aside for twenty-four hours. The 
number on the scale on the glass tube which marks the height 
of the coagulated albumin represents the percentage of albumin 
in the urine. To insure accuracy the tube should stand in a 
temperature of about 15° C. (60° F.), and the specific gravity 
of the urine should be reduced to between 1006 and 1008 by 
dilution, the amount of dilution being taken into consideration 
in calculating the percentage of albumin. 

Albumosuria. — Albumose appears in the urine as a part 
product of proteid metabolism. It is a pre-peptone which ap- 



ALBUMIN. 



199 



pears in the blood and which can be produced by artificial diges- 
tion. Pathologically, its continued and marked presence in 
the urine usually denotes multiple myelomata. It has also 
occasionally been noted in syphilis, pneumonia, peritonitis, and 
cholera. 




Fig. 24.— Esbach Albuminometer. 

At end of 24-hour test albumin per cent, reading a 
fraction less than 4 per cent. 



The Test. — Add concentrated nitric acid to the urine, agi- 
tate and allow to stand in a cool place. A heavy precipitate will 
occur which, upon heating, disappears, only to reappear on cool- 
ing. At the same time the mixture gradually assumes an 
intense yellow color. 

Albumin of Bence Jones. — This substance has been repeat- 
edly found in cases of multiple myelitis. Its exact nature is 



200 THE URINE. 

not definitely known, but it is generally regarded as an albumose. 
Its reaction to heat and nitric acid is similar to that of albn- 
mose. The test for its positive identification is complicated and 
difficult, and will hardly be required of the clinical laboratory 
worker. 

Nucleo-Albumin. — Nucleo-albumin, beyond a faint trace, 
will give the same reactions as true blood-albumins. Its pres- 
ence may be inferred when there is an albumin reaction in a 
specimen which shows an absence of renal elements with an 
excess of epithelia of bladder and urethral type. 

Test. — To dilute acid urine add an excess of acetic acid, 
stand aside, and gradually a faint cloud will appear. 

Differential Test. — If the preceding test gives a positive 
reaction, take a fresh portion of the same specimen, and first acid 
one-sixth volume of a saturated sodium chlorid solution, and 
then some dilute acetic acid. The presence of sodium chlorid 
will prevent the precipitation of nucleo-albumin in the last test. 

Fibrin, when present in the urine, usually occurs in suf- 
ficient amounts to form clots, which are easily distinguishable. 
When the amount is small it may become necessary to determine 
the presence of this substance by chemical means. 

Test for Serum-Globulin. — Add to some urine contained 
in a small beaker sufficient ammonia water to produce a slightly 
alkaline reaction; a cloud of phosphates will appear and will 
settle to the bottom. As soon as a layer of clear urine appears, 
decant this into a test-tube. Mix this clear urine with an equal 
quantity of a clear saturated solution of ammonium sulphate. 
If a flocculent precipitate appears in a few minutes the albumin 
present is serum-globulin. 

Test for Serum-Albumin. — Take the final solution ob- 
tained in the foregoing test and filter it free from serum-globu- 
lin. Heat the filtrate to boiling, and add one-tenth volume of 
strong nitric acid. If a precipitate now appears the urine con- 
tains serum-albumin. 

Proteose. — Mix about 50 cubic centimeters of urine con- 
tained in a beaker with 5 cubic centimeters of strong nitric 
acid. Heat the solution rapidly to about 80° C. If a pre- 
cipitate appears, prepare a hot filter-paper by pouring boiling 
water through it in a funnel. Now filter the hot mixture of 



GLUCOSE. 201 

urine and nitric acid. Add to the clear, hot filtrate its own 
volume of a clear, saturated solution of sodium chlorid. Allow 
the mixture to cool. If any precipitate appears in this cooled 
mixture then proteose was present in the specimen of urine 
under examination. 

GLUCOSE. 

General Considerations. — A trace of sugar can at times be 
detected in apparently normal urine from healthy subjects by 
special tests of great delicacy. The ordinary tests employed in 
clinical medicine, however, do not react to these minute amounts 
of sugar, hence the findings of glucose by the tests about to be 
outlined must, in every case, be considered pathologic. 

The reducing powder of normal urine is, in part, due to the 
activity of uric acid and creatinin, and is equal to about a 0.1- 
per-cent. solution of glucose. 

Under pathologic conditions the reducing power of urine 
may be enormously increased, due to the abnormal presence of 
glucose. This may be present up to as much as fifty grains to 
the ounce (10 per cent.). 

The detection of sugar in the urine is based upon a knowl- 
edge of the following properties of glucose : 

1. In hot alkaline solutions, reduces the oxides of copper 
and bismuth. 

2. With brewer's yeast, ferments, forming alcohol and car- 
bon clioxid. 

3. Enters into chemical combination with phenylhydrazin 
to form characteristic insoluble crystals of glucosazone. 

4. Deflects polarized light to the right. 

In the application of all tests but the fermentation-test, it 
is necessary to use urine free from albumin. If this be present 
it must first be removed by the addition of dilute acetic acid and 
boiling to cause precipitation. This is then filtered out, and the 
filtrate used in the tests for sugar. 

Reduction Tests. — Method of Fehling 3 (for preparation 
of reagent see Appendix) : The sample of urine to be tested is 
first examined carefully for the presence of albumin. If this be 



3 The suggestions forthe performance of this test are taken from a communication 
by C. W. Louis Hacker, M.D., appearing in the Jour. Amer. Med. Asso., page 252, 
Jan. 25. 1908. 



202 THE URINE. 

found it must be removed by heat and acetic acid, and the filtrate 
used for the test. About -± cubic centimeters of freshly pre- 
pared Fehling^s solution are diluted with three parts of water, 
and brought just to the boiling point in a clean test-tube. 
Immediately two drops of proteid free urine are added, and the 
tube shaken vigorously. If the urine contains more than 2 per 
cent, of sugar, a yellowish-red precipitate immediately appears, 
changing rapidly to a brownish-red or bright red, which settles 
out slowly, leaving a greenish-blue supernatant fluid. If the 
reaction be positive, it is better to roughly dilute the urine with 
two or four parts of water, and to perform the test again. 

With the strengths of dextrose from % to 2 per cent., two 
drops of urine, as stated above, will cause a characteristic pre- 
cipitation of red suboxid of copper, about which there can be 
no doubt. With smaller percentages the change may come slowly, 
but even then heat need not be applied after the addition of the 
urine. Here little change may be seen in the Fehling's solution 
by transmitted light, but by reflected light a brownish-red tint 
is evident, which finally develops into a decided light-red pre- 
cipitate, gradually settling out and leaving a clear, blue, super- 
natant fluid. The pentoses, lactose, and even maltose, by this 
method, do not give the characteristic reaction. Urines showing 
this reaction always give a reaction with Xylander's solution 
(see next page). 

If no change occurs in the Fehling's solution after the addi- 
tion of the two drops as above, the solution may then be warmed 
just to the boiling point, and from two to four drops of urine 
added and the mixture observed. This should be repeated until 
in all twenty drops of urine have been added. If no change 
occurs at this point, the urine does not contain dextrose to the 
extent of 0.5 per cent. 

Cautions. — The disadvantages of using large amounts of 
urine (an equal or even one-half volume of the Fehling's solu- 
tion) are two-fold. In the first place, the strongly alkaline cop- 
per solution with urine of high specific gravity throws down a 
more or less voluminous precipitate of cupric phosphate, which 
takes out the blue color of the solution, and changes it to a dirty, 
dark green. In the second place, the alkaline solution, acting on 
the ammonium salts in the urine, liberates sufficient ammonia to 



GLUCOSE. 203 

hold in solution small amounts of the red suboxide of copper, 
obviously interfering with the detection of traces of dextrose. 
Very frequently under these conditions, and especially if the 
solution has been boiled, even for a few seconds after the addi- 
tion of the urine, the greenish solution slowly takes on a dirty, 
yellow-brown color, then become turbid, and, eventually, on 
standing, there separates out a finely divided opalescent, green- 
ish-yellow precipitate, which only settles out completely after the 
tube has been allowed to stand over night. In certain cases this 
reaction does not begin until a few seconds have elapsed after 
the addition of the urine. Such a reaction may be termed a 
pseudo-reaction. 

Fallacies. — Neglect of the above precautions will result in 
the occurrence of the pseudo-reaction in about 5 per cent, of nor- 
mal urines examined. 

The following substances, if present in the urine in more 
than normal amounts, will result in partial reduction of Feh- 
ling^s solution. Uric acid, creatinin, hippuric acid, mucin, 
hypoxanthin, and alkapton. Also the presence of the oxidation 
products of the following drugs : The alkaloids, arsenic, carbolic 
acid, and hexamethylamin (urotropin). The latter substance 
does not ordinarily reduce bismuth. 

Bottger's Bismuth Test. — Urine containing coagulable 
proteids must first have them removed by heat and acetic acid. 
Mix in a clean test-tube equal volumes of urine and liquor po- 
tassii, and add a few grains of bismuth subnitrate and boil. In 
the presence of glucose the white pow r der in the bottom of the 
tube will gradually become gray, brown, or black. 

ISTylander's Bismuth Test. — Nylander modifies the pre- 
ceding test by substituting for the subnitrate a special reagent 
containing bismuth (for reagent see Appendix). Albumin, if 
present in the urine, must be removed in the usual way. To 
10 cubic centimeters of Nylander's reagent, add 1 cubic centi- 
meter of urine and bring to the boiling point. A brown or 
black discoloration of the liquid denotes sugar. 

Fallacies of bismuth tests. A slight reduction of bismuth 
may be caused by turpentine, eucalyptus, or the ingestion of large 
amounts of quinine. Albumin and sulphur compounds in urine 
produce a black precipitate. 



204 THE URINE. 

Delicacy, — These tests show the presence of glucose in 
0.025-per-cent. solution. 

Trommer's Test. — To half inch of urine in a test-tube, add 
one inch of liquor potassii and a few drops of a 10-per-cent. solu- 
tion of copper sulphate sufficient to cause blue discoloration with 
slight cloudiness. Heat this mixture to the first signs of boil- 
ing. If sugar be present the copper will be reduced to yellow- or 
red-oxid. Slight precipitations, which may occur after pro- 
longed boiling, are no proof of sugar. 

Delicacy. — This test, if carefully performed, will demon- 
strate the presence of sugar in 0.025 per cent, solution. 

Modifications of Trommer's Test (Simrock). — Mix 
equal parts of the Simrock reagent (see Appendix) and urine 
in a test-tube and boil ; the presence of glucose will be indicated 
by a yellowish or red discoloration and precipitate. 

Phexylhydrazix Test. — This test depends upon the pro- 
duction of characteristic crystals of phenylglucosazone by combi- 
nation of glucose and phenylhydrazin in hot solution, and their 
recognition with the aid of a microscope. 

The Test. — Fill a beaker half full of water and warm upon 
the wire-gauze over a low Bunsen flame. Place a test-tube, con- 
taining the urine, which has been acidulated with a few drops of 
dilute acetic acid, in the beaker. While waiting for this to 
warm prepare the following solution (this must be made fresh, 
as it does not keep well) : Weigh out one gram of phenylhy- 
drazin hydrochlorid, and two grams of sodium acetate. Mix the 
salts and dissolve in 10 cubic centimeters of distilled water 
acidulated with two drops of dilute acetic acid. Filter to clarify. 
After the water in the beaker has boiled for five minutes, observe 
the urine in the tube; if it has become turbid, filter. To 10 
cubic centimeters of hot urine in a clean test-tube, add 5 cubic 
centimeters of the filtered reagent, replace in a beaker of boiling 
water, and continue boiling for one hour. Then cool quickly by 
immersing the tube in cold water. If sugar was present in the 
urine a crystalline precipitate will appear. This should be taken 
up in a pipette and transferred to a microscope slide for exami- 
nation. The characteristic crystals of phenylglucosazone are 
fine, faintly yellow needles arranged in the forms of fans, 
rosettes, and sheaves. 



QUANTITATIVE ESTIMATION OF GLUCOSE. 205 

Fallacy. — Levulose, if present in the urine will produce 
similar crystals; these can be differentiated by a comparison of- 
the fusing points of the two compounds. 

Phenylglucosazone fuses at 20-1° C. 

Phenyllevulosazone fuses at 150° C. 

Delicacy. — Glucose forms characteristic crystals when pres- 
ent in 0.025 per cent, solution. 

QUANTITATIVE ESTIMATION OF GLUCOSE. 

Volumetric Determination by Fehling's. — Measure into a 
beaker or porcelain dish 10 cubic centimeters of accurately pre- 
pared Fehling^s solution, and 40 cubic centimeters of distilled 
water. Heat the mixture over wire gauze to boiling. Prepare 
a dilute solution of suspected urine by adding nine parts of 
distilled water to one part of urine. This mixture is placed in 
a burette. Xow to the boiling solution add the dilute urine, a 
few drops at a time, from the burette. Continue adding, boil- 
ing, and stirring until the blue color of the Fehling^s solution 
completely disappears when view r ed by transmitted light. Note 
accurately the number of cubic centimeters of urine used. 

Fehling's solution is a standard solution of copper sulphate, 
10 cubic centimeters of which is exactly decolorized by 0.05 
gram of glucose. 

Example. — Suppose 8 cubic centimeters of diluted urine 
were used in decomposing 10 cubic centimeters of Fehling's 
solution. Then, of undiluted urine, 0.8 cubic centimeter was 
required. This amount then contained 0.05 gram (10 x 0.005) 
glucose. To calculate the percentage of sugar in the sample 
of urine: 0.8: 0.05:: 100: X, which, in this case, "X" equals 
6.25 per cent. To calculate the grams of glucose voided in 
twenty-four hours: If 1500 cubic centimeters of urine were 
voided in twenty-four hours, then 0.8 : 0.05 : : 1500 : X, which 
equals 93.75 grams glucose. 

Caution. — The dilute urine should be added very slowly 
with frequent boilings and examinations by transmitted light to 
avoid passing the end reaction. 

Robert's Differential Density Method. — This method de- 
pends for its result upon the alteration in density occuriing from 
the fermentation of saccharine urine. 



206 THE URINE. 

The Test. — Mix thoroughly about 120 cubic centimeters 
of urine with half a cake of compressed yeast. Take and 
record accurately the specific gravity of this mixture. Set aside 
in a warm place (thermostat preferred) for twenty-four hours, 
and at the expiration of this time again take the specific gravity 
and subtract the second reading from the first. Each degree 
of the remainder (showing density lost) represents approxi- 
mately one grain of sugar to the ounce. To obtain the per- 
centage of sugar in the specimen, multiply the degrees of 
density lost through fermentation by the factor 0.219. 

Example. — Specific gravity before fermentation 1041 

Specific gravity after fermentation 1023 

Degrees of density lost 18 

This is approximately the grains of glucose in each ounce of urine. 
18x0.219 equals 3.94% glucose. 

This test is easily performed, and is conclusrve evidence of 
the presence of glucose. It is, however, not strictly accurate, and 
is not to be relied upon when the amount of glucose present is 
less than 0.5 per cent. 

FERMENTATION SACCHARIMETER. 

None of the substances found in the urine, which give the 
reduction reactions resembling glucose, are susceptible of alco- 
holic fermentation. This fact establishes the accuracy of the 
test. The test itself depends upon the fact that when sugar- 
containing urine is mixed with a quantity of brewer's yeast and 
kept in a warm place for twenty-four hours, it will ferment, 
giving off bubbles of C0 2 , and at the same time forming 
alcohol. 

By measuring the amount of carbon dioxid gas, formed 
during this fermentation-process, we are enabled to estimate the 
percentage and amount of sugar contained in the specimen un- 
der examination. A specially shaped and graduated tube (see 
Fig. 25) is usually employed in this test. It is known as the 
Einhorn saccharimeter. The upright tube is graduated from its 
closed upper end downward from 0.1 to 1.0 per cent. It will not 
estimate sugar in amounts less than 0.1 per cent., and owing to 
the varying solubility of the various gases of the urine, depend- 



FERMENTATION SACCHARIMETER. 



207 



ing on differing reactions and the density, it cannot be regarded 
as absolutely accurate. 

The Test. — Urines containing less than 5 per cent, of 
glucose must be diluted five times with water. Urines contain- 
ing more than 5 per cent, must be diluted ten times before pro- 
ceeding with the test. 




a b 

Fig. 25.— Einhorn Saccharimeter During Performance of Test. 
A, Showing 2>£ Per Cent. Sugar. B, Control with Normal Urine. 



Mix in a large test-tube about 20 cubic centimeters of 
properly diluted urine with one-twelfth of a cake of fresh, 
compressed yeast. Completely fill the upright tube of the 
saccharimeter with this mixture, so as to exclude all air from 
the graduated tube. Fill a second saccharimeter with yeast 
dissolved in distilled water or with urine known to be free from 



208 THE URINE, 

sugar. These tubes are to be kept in a warm place and 
allowed to remain undisturbed for from eighteen to twenty- 
four hours. By this time the sugar-containing urine will be 
found to have been displaced by gas in the vertical tube. The 
percentage corresponding to the level of the fluid will represent 
the percentage of sugar in the diluted urine. This figure mul- 
tiplied by the dilution, will represent the percentage of sugar 
in the specimen under examination. 

Cautions. — The urine must be faintly acid, and must be 
so diluted that the specific gravity will be less than 1008, and 
when diluted must contain less than 1 per cent, of glucose. If 
any gas is liberated by the yeast or fluid in the control-test, this 
amount must be subtracted from that indicated in the test-mix- 
ture before computing the percentage. 

The internal administration of mercuric ehlorid, iodoform, 
salicylic acid, hexamethylamin, quinine, and other antiseptic 
drugs will inhibit fermentation, and therefore must be excluded 
before testing. 

PURDY'S QUANTITATIVE METHOD. 

Into a beaker or boiling flask of 250 cubic centimeters 
capacity put 35 cubic centimeters of Purdy's reagent (see 
Appendix) and 70 cubic centimeters of distilled water. Boil 
steadily over a wire gauze, and add urine slowly from a burette 
until the blue color begins to fade; now proceed cautiously, and 
after the addition of each drop wait for a few seconds to see if 
the end reaction is complete. Continue adding until the solu- 
tion of the reagent is colorless and transparent. To obtain this 
result the total amount of urine employed must have contained 
0.02 gram of glucose. 

Example. — Suppose the amount of urine employed to com- 
pletely decolorize the reagent was 4.5 cubic centimeters, then 
4.5 cubic centimeters of urine contained 0.02 gram of glucose; 
or 1 cubic centimeter of urine contains 0.0044 gram of glucose. 
This multiplied by 100 will give the percentage of glucose which 
is equal to 0.41. If the color of the reagent is changed by less 
than 4 cubic centimeters of urine, it is best to dilute the urine 
with one or two parts of water, and then multiply by this factor 
to obtain the final result. 



CAMMIDGE REACTION. 209 

This method is rapid, the technic is simple, and the end- 
reaction definite and sharp. The reagent is stable. 

CAMMIDGE REACTION.* 

This reaction is based upon the presence of certain, at pres- 
ent unknown, substances, which occur in the urine in pancreatic 
disease associated with fat necrosis, the detection of which is 
based upon some special reactions in which phenylhydrazin hy- 
drochlorid plays an active part. 

A-Keaction. — The specimen of urine to be examined is 
filtered, and 10 cubic centimeters of the filtrate are placed in a 
small flask; 1 cubic centimeter of hydrochloric acid (specific 
gravity 1.16) is added, and a funnel placed in the neck of the 
flask to act as a condenser. The flask is then set up on a sand- 
bath and gently boiled for eight minutes; then, after rapid 
cooling in running water, the contents are filtered. 

To 5 cubic centimeters of this filtrate add 5 cubic centi- 
meters of distilled water. The excess of acid is now neutralized 
by the slow addition of four grams of lead carbonate; then, 
after standing for a few r minutes to allow completion of the 
reaction, the urine is filtered through a wet filter and the flask 
washed out with 5 cubic centimeters of distilled water through 
the filter. To the clear filtrate now aclcl two grams of pul- 
verized sodium acetate, and 0.75 gram phenylhydrazin hyclro- 
chlorid. This mixture is now boiled for from three to four 
minutes on the sand-bath. The hot fluid is then immediately 
poured into a test-tube and allowed to cool undisturbed. After 
the lapse of a variable period, depending on the severity of the 
case, of from one to twenty-four hours, a more or less flocculent 
precipitate or yellowish deposit accumulates in the bottom of 
the tube. This precipitate, when examined under the micro- 
scope, is found to consist of sheaves and rosettes of golden 
yellow crystals. 

As a necessary preliminary to this test, sugar, if any be 
present in the specimen, must be removed by fermentation, and 
the alcohol formed driven off by heat. Albumin should be re- 
moved by the addition of dilute acetic acid, heating and filtering. 



4 See "Diseases of Pancreas," Robson and Cammidgfe, 1907, for further informa- 
tion. 



210 THE URINE. 

When the A-Keaction is positive then the B-Beaction must 
be made. 

B-Beaction. — Twenty cubic centimeters of urine are 
filtered and mixed with 10 cubic centimeters of a saturated 
bichlorid of mercury solution. After filtering, the filtrate is 
allowed to stand for a few minutes, and then to 10 cubic centi- 
meters of this filtered mixture add 1 cubic centimeter of 
concentrated hydrochloric acid. This is then placed in a small 
flask and transferred to a sand-bath, and allowed to boil for 
ten minutes. Subsequently to 5 cubic centimeters of this mix- 
ture add 10 cubic centimeters of distilled water. After cooling, 
this is neturalized with four grams of lead carbonate. The 
remaining stages are the same as in the A-Eeaction. 

The B-Eeaction is a differential one based upon the ob- 
servation of Cammidge, that in inflammatory conditions of the 
pancreas the crystals formed in the A-Eeaction are destroyed by 
the mercuric bichloride. 

A large number of observations showed Cammidge that the 
crystals in malignant disease are broad and coarse, while in 
inflammatory processes they are smooth and slender. 

The solubility of the crystals in 33 per cent, sulphuric acid 
also differs according to the nature of the affection. 

The crystals obtained from a case of acute pancreatitis are 
soluble in from one-quarter to one-half minute ; in chronic pan- 
creatitis in from one-half to two minutes ; in carcinoma in from 
three to five minutes or longer. 

Negative results with these reactions have been obtained by 
Cammidge, 5 and also by Eobson, 6 in normal urines, in jaundice, 
in gall-stones without pancreatitis, in gastric ulcer, and in a 
number of other conditions. 

In support of the practical value of these reactions it may 
be stated that Felix Eichler 7 has lately produced acute pancrea- 
titis in dogs, and in the urine of each he found the character- 
istic crystals, while the reaction applied to normal control dogs 
it was consistently negative. 



5 Cammidgre: Lancet, March 19, 1904. 

6 Mayo Robson : Laucet, page 773, 1904. 

7 F. Eichler: Berlin, klin, Wochenschrift. 






CLINICAL SIGNIFICANCE OF GLYCOSURIA. 211 

POLARIMETRIC METHOD. 

Because of its ability to turn the ray of polarized light to 
the right, glucose is called dextrose. The accurate determination 
of glucose in urine may be made by those possessing polari- 
scopes. The degree of dextro-rotation can be read on a gradu- 
ated scale and calculated as percentage or grams of glucose. 

Since albumin in solution deflects the polarized ray to the 
right, this must first be removed by acidulating, boiling, and 
filtering. To make an accurate estimation the urine must be 
first decolorized by shaking with a piece of lead acetate and 
filtering. 

Delicacy. — Instruments vary in delicacy. Usually dis- 
tinct dextro-rotation may be detected when glucose is present in 
0.025 per cent. 

Fallacy. — Maltose, which rarely occurs in the urine, if 
present, will deflect the ray even more than will glucose. 

(For description of technic see larger works on clinical 
chemistry or polarimetry.) 



CLINICAL SIGNIFICANCE OF GLYCOSURIA. 

The presence of glucose in the urine is pathologic. If the 
amount be abundant and persists, associated with copious water- 
drinking and eating, while the patient is at the same time 
emaciating, it is indicative of diabetes mellitus. The urine in 
this condition is usually pale with a fruity odor, and while over- 
abundant, has a high specific gravity. As much as 10,000 cubic 
centimeters of urine may be excreted in twenty-four hours, at the 
same time the specific gravity may be maintained at 1050. 

Temporary Glycosuria. — The appearance of small amounts > 
of sugar may occur transiently in the urine after excessive inges- 
tion of sugar; and reducing substances, such as glycuronic acid, 
may give a spurious positive reaction. Copper reduction by such 
substances is not corroborated by either fermentation or the 
phenylhydrazin tests. 

Glycosuria also may accompany certain diseases of the 
brain, spinal cord, and pancreas, or be a transitory accompani- 
ment of phthisis, pneumonia, cirrhosis, or cholera. 



212 THE URINE. 

LACTOSEURIA. 

Milk sugar occurs occasionally in the urine of nursing 
women, particularly toward the end of lactation. It will reduce 
Fehling^s and Xylander's solutions, but returns a negative result 
with the phenylhydrazin test. It ferments very slowly. 

MALTOSEURIA. 

Maltose has been found in the urine of diabetic patients, 
and when present reduces copper and ferments with yeast. Its 
occurrence is sufficiently rare to be discounted in the clinical 
laboratory. 

PENTOSEURIA. 

Diabetics and morphine habitues occasionally show in the 
urine traces of pentoses. These will reduce copper, but will not 
ferment. 

Newmann's Orcin Test. — This test may be employed to dis- 
tinguish between the pentoses, the glycuronates, and dextrose. 

Techxic. — Three cubic centimeters of urine are treated 
with ten drops of a 5-per-cent. alcoholic orcin solution and 10 
cubic centimeters of glacial acetic acid. The mixture is now 
brought to the boiling point and allowed to cool. After cooling 
concentrated sulphuric acid is added, drop by drop, and shaken 
until about twenty drops have been added. In the presence of 
pentoses the color becomes olive-green, glycuronates turn the 
solution violet, and dextrose gives a carmine color. 

ACETONE. 

Usually in advanced diabetics, sometimes in fever or in 
.perfect health, particularly in patients whose diet is rich in pro- 
teids, the urine may have an ethereal odor, and will give posi- 
tive reactions to tests for acetone. 

In the course of a case of diabetes a decline in the percent- 
age of sugar, accompanied by lowered specific gravity, but with- 
out corresponding improvement in the general symptoms, may 
indicate impending coma. At this time acetone will be found 
in the urine in increasing amounts, with or without diacetic acid 
and oxvbutvric acid. 



ACETONE. 213 

Ethylene-Diamin-Hydrate Test. — This is the simplest, and 
is probably the most satisfactory for clinical purposes. 

Test. — Place in a clean test-tube about 5 cubic centimeters 
of distilled water, and to this add about one grain of sodium 
nitro-prusside : shake to effect solution. Add to this an equal 
amount of suspected urine and thoroughly mix. Gently overlie 
the mixture with a few drops of a 10-per-cent. solution of 
ethylene-diamin-hydrate. A pink or ruby-red color at the zone 
of contact denotes the presence of acetone. A faint white cloud, 
which appears on the addition of the reagent, does not denote 
acetone. The solution of sodium nitro-prusside must be freshly 
prepared. 

Legai/s Test. — To 5 cubic centimeters of urine in a test- 
tube, add a few drops of a freshly prepared solution of sodium 
nitro-prusside and a few drops of sodium hydrate solution. A 
red color will appear in all urine, which soon changes to yellow. 
If this is now overlaid wdth strong acetic acid a change in color 
at the line of contact from yellow to carmine or purplish-red 
indicates acetone. 

DIACETIC ACID. 

Coincident with the appearance of acetone in diabetic urine, 
we may have diacetic acid. 

Test. — To 5 cubic centimeters of urine in a test-tube, add 
two drops of ferric chlorid solution; if a precipitate appears, 
filter and add more ferric chlorid to the filtrate. The presence 
of diacetic acid will be indicated by a portwine discoloration. 

FtfZ/aci/.— Antipyrine, salicylic acid, and coal-tar derivatives 
will give a similar reaction if they have recently been ingested 
by the patient. 

OXYBUTYRIC ACID. 

This substance cannot be readily detected by chemical means 
unless a long and difficult preliminary process is resorted to in 
order to separate it from the other urinary constituents. These 
methods will be found in larger works on clinical and physio- 
logic chemistry. 



214 THE URINE. 

BILE PIGMENTS. 

General Considerations. — Bile pigments are never found in 
the urine under normal circumstances. As a rule the freshly 
voided urine only contains the oxidized derivative, bilirubin. If 
a cystitis should exist, then subsequent oxidation products, 
biliverdin, bilifuscin, biliprasin, and bilihumin may also be 
encountered. 

Bile-containing urines present a more or less characteristic 
appearance ; their color may vary from a bright golden-yellow to 
a dark greenish-brown. On microscopic examination it is usual 
to find the morphologic elements of the urine stained yellow or 
reddish-brown. The same color may be imparted to the foam 
on shaking. 

Gmellin's Test for Bile-Pigments. — Put some yellow nitric 
acid in a test-tube and gently overlay it with the suspected urine. 
In the presence of bile-pigment a play of colors will be observed 
at the zone of contact. Green will be seen nearest the urine and 
orange in the upper part of the nitric acid. This test is exceed- 
ingly sensitive, indicating the presence of bile-pigment in a dilu- 
tion of 1 to 80,000. 

Nitric Acid Paper Test. — Moisten a piece of white filter or 
blotting paper with the suspected urine, and place it on a 
glazed tile or slab of porcelain. Allow one or two drops of 
commercial nitric acid to fall into the center of the wet paper. 
In the presence of bile-pigment concentric rings of blue, violet, 
green, and yellow will appear. A slight red reaction cannot be 
considered positive, as it may be produced by other substances 
than bile-pigment. 

Import. — When bile is not freely and normally discharged 
from the bile-passages, the coloring matter from the retained 
bile is absorbed by the lymphatics, the various body tissues be- 
come stained with it, and it is partly eliminated in the urine. 

BILE ACIDS. 

The presence of bile-acids in the urine has the same clinical 
significance as the presence of biliary pigment. The tests for the 
acids are fraught with considerable difficulty, and do not com- 
pensate in their significance for the time and energy expended. 



HEMATURIA. 215 

It will, therefore, usually be found sufficient to determine the 
presence of bile-pigments. 

UROBILIN. 

General Considerations. — Urobilin is closely allied to the 
normal urochrome, but is an abnormal product, probably result- 
ing from the action of reducing agents upon the normal urinary 
pigment. It is said to be identical with the stercobilin of the 
feces, and its behavior to chemical reagents can be tested by 
testing an alcoholic extract of feces. 

Test for Urobilin. — (a) Mix some urine in a test-tube with 
an equal amount of a 10-per-cent. solution of zinc acetate. Filter 
off the precipitate, and the filtrate, in the presence of a demon- 
strable quantity of urobilin, will show a beautiful greenish 
fluorescence. With the polariscope the urobilin in this solution 
gives a distinct absorption spectrum. 

(b) Fill a clean test-tube three quarters full of urine, and 
add one drop of strong hydrochloric acid ; to this add about one- 
sixth volume of amylic alcohol, and after shaking slowly eight 
or ten times allow to separate by standing. Pour off the super- 
natant fluid and add to it thrice its volume of alcohol. Prepare 
separately a 5-per-cent. solution of zinc ehlorid, and add one 
drop of this to the alcoholic extract of the urine; finally add 
one drop of ammonium hydroxid. Zinc hydrate will be pre- 
cipitated, which should be filtered off. In the presence of 
urobilin the filtrate will present a beautiful green fluorescence. 

Significance. — Urobilin has been detected in cases of hepatic 
cirrhosis, malarial anemia, carcinoma, Addison's disease, and 
pancreatic disease with acholic stools. 

HEMATURIA. 

General Considerations. — Blood may gain access to the 
urinary tract in many ways, and appear in the urine in varying 
amounts, from the smallest trace, demonstrable only by chem- 
ical means, to sufficient to produce bloody urine with clots. 

Blood from the kidney is usually well mixed with the urine, 
to which it imparts a brownish smoky hue. Under the micro- 
scope tube-casts of blood-cells may be found. 



216 THE URINE. 

Blood from the ureter may be well-mixed with the urine, 
or may appear in characteristic worm-like clots. 

Blood from the bladder may, or may not, show irregular 
clots. When recently shed it imparts a bright red color to the 
urine; it is frequently accompanied by much mucus and large, 
flat epithelial cells in great numbers. 

Blood from the prostate, examined by the three-glass test, 
appears in the first and third glasses only. 

Blood from the urethra appears in the first glass, and is 
frequently clotted. 

Urine may show the presence of blood in the absence of 
any demonstrable lesion of the genito-urinary tract. It has 
been noted after the ingestion of strawberries, gooseberries, or 
a large amount of rhubarb. 

The blood may be contaminated from menstrual discharge. 
This possibility should always be borne in mind, and false con- 
clusions guarded against. 

Microscopic Appearance. — The most convenient method of 
demonstrating blood in the urine is by microscopic examination 
of the centrifugatecl or sedimentated sample. The corpuscles 
in ordinary acid urine maintain their characteristic bi-concave 
shape for a number of days, if decomposition and putrefaction 
are prevented. If the urine is of high specific gravity, the cells 
rapidly become crenated. On the other hand, if the urine be 
alkaline or becomes so after voiding, the corpuscles will appear 
swollen, shriveled, or shadowy. Urine containing more than a 
trace of blood is albuminous. 

Test for Occult Blood in the Urine. — To 10 cubic centi- 
meters of urine in a large test-tube, add about twice as much 
ether and agitate thoroughly by pouring from one test-tube to 
another several times. To this add a few grains of powdered 
gum guaiac, and again agitate. Xext add 5 cubic centimeters 
of glacial acetic acid (99.4 per cent.) and again agitate; allow 
this to settle and then pour off the supernatant liquid and 
divide equally between two test-tubes; set one aside for a con- 
trol, and to the other add about 2 cubic centimeters of fresh 
hydrogen dioxid from a pipette, making an effort to have it 
settle to the bottom as a distinct laver. If a bluish discolora- 



PYURIA. 217 

tion appears either at the zone 6i contact or throughoul the 
mixture, the presence of blood is demonstrated. 



HEMOGLOBINURIA. 

This term signifies the presence of hemoglobin in the urine 
free from blood-corpuscles. Besides the foregoing test it may 
be demonstrated by means of the spectroscope. Faintly-acid 
urine containing traces of hemoglobin will give two character- 
istic absorption bands of oxyhemoglobin. By the addition of a 
minute quantity of ammonium sulphicl, the spectrum is changed 
to that of reduced hemoglobin. 

Hemoglobinuria occurs in a variety of conditions : scurvy, 
pyemia, purpura, typhus fever, poisoning from arsenic, phos- 
phorus, carbolic acid, chloral, and potassium chlorate. There 
has also been noted a periodic form of obscure origin. 

PYURIA. 

Pus being an albuminous fluid, will cause urine containing 
more than a minute quantity to respond to the tests for albu- 
min. Urine containing much pus is turbid, and deposits on 
standing a white or greenish-white sediment which is insoluble 
in heat and in dilute mineral acids. The addition of hydrogen 
dioxid produces rapid effervescence. 

Microscopic examination of pus-containing urine will re- 
veal the characteristic pus corpuscles. These are spheric, granu- 
lar, and highly retractile. The nucleus is usually obscured or 
disintegrated. 

Donne's Test for Purulent Sediment. — To the suspected 
sediment add a small piece of caustic potash, and stir with a 
glass rod. Pus will become thick, tough, and gelatinous, while 
mucus will become flaky and thin. 

Significance of Pyuria. — The presence of pus in the 
urine indicates the presence of an inflammatory process in the 
genito-urinary tract, the location of which can, in some measure, 
be determined by the character of the associated epithelial cells 
(see page 218). 



218 THE URINE. 

EPITHELIA 

The epithelial cells found in urine may come from any part 
of the genito-urinary tract. Their forms vary greatly, and 
with a knowledge of the characteristic cells belonging to the 
different regions, it is possible, with more or less certainty, to 
determine their origin by their appearance. The puss cell may 
be taken as the standard of size. 

Cells from the tubules of the kidney are round and about 
one-third larger than a pus cell. 

From the pelvis of the kidney, twice the size of a pus cell 
and cuboidal or pear-shaped. 

From the ureter, round and slightly smaller than from the 
pelvis. 

From the bladder, flat and square. These are the largest 
cells encountered, except those from the vagina. 

From the urethra, smaller than from the bladder ; they may 
be cuboidal or columnar. All epithelial cells are granular and 
contain a relatively small nucleus. 

TUBE CASTS. 

General Considerations. — In the presence of albuminuria or 
hematuria, microscopic examination of the urinary sediment will, 
as a rule, reveal the presence of tube casts. Occasionally some 
varieties of casts will be found in urine which show neither 
albumin or blood. 

The Sediment. — In order to obtain a sediment for micro- 
scopic examination, some method of precipitating and of con- 
centrating this precipitate is necessary for a conclusive exami- 
nation. A centrifuge (Figs. 26, 27 and 28) is the most rapid 
as well as the safest method to employ. This process only 
requires from 15 to 20 cubic centimeters of urine, and can be 
accomplished in a few minutes. After centrifugation a part 
of the supernatant urine is poured off and then, with the aid 
of a pipette with a small point, a drop or two is drawn up and 
transferred to a clean microscope slide for examination. 

The method of employing a conical glass may be used if a 
centrifuge is not at hand. Sedimentation by this means re- 
quires a number of hours, and, if care is not taken to prevent 



TUBE CASTS. 



219 



bacterial contamination and if the specimen is not kept in a cool 
place, the specimen may decompose and the morphologic ele- 
ments be destroyed. 

The sedimentation glass is valuable for roughly estimating 
the amount of gross sedimentation in phosphaturia, pyuria, etc. 
(Fig. 29.) 




Fig. 26.— Water-power Centrifuge. 





Fig. 27.— Electric-power Centrifuge. 



Fig. 28.— Hand-power Centrifuge. 



Varieties of Casts. — Hyaline casts are almost transparent 
and appear as ground glass. They have a delicate but definite 
outline, and are quite friable. 

Strong illumination of the field may obscure them entirely, 
as they are very delicate in outline and structure. They are best 
seen with the plane side of the reflector, and side illumination 
which should not be too bright. 

The particular significance of hyaline casts is not yet posi- 



220 THE URINE. 

tively settled. They occur in advanced grades of nephritis and 
again in transient albuminurias, and even in the absence of 
demonstrable albumin; they are frequently seen during the 
course of fevers, particularly typhoid fever. 

Granular Casts. — These are more opaque than the hya- 
line, and are, therefore, more easily seen. The granules are 
numerous, and upon close examination will be found to permeate 
the matrix of which the cast is partly composed. This will serve 
to differentiate them from the pseudo-granular casts, which are 
merely hyaline casts to which granular debris has become at- 
tached during centrifugation or sedimentation. These latter 




B C 

Fig. 29.— Sedimentation Glasses. 

A Clear. B, Slightly cloudy with beginning precipitation. 
C, Sedimentation complete. 

are of no more significance than hyaline casts. Granular casts 
are probably composed of a hyaline matrix which has undergone 
degeneration. They frequently show fragmented cells and fat 
globules in their structure. The continued occurrence of hya- 
line and granular casts in the presence. of a permanent albu- 
minuria, usually denotes chronic interstitial nephritis. 

Hyalo-granular Casts. — Hyaline casts are occasionally 
found, parts of which are distinctly granular throughout their 
substance. They apparently represent a stage of cast-formation 
between the hyaline and granular varieties, and their significance 
is probably the same as hyaline casts. 

Epithelial Casts. — These casts are composed of renal 



CYLINDROIDS. 221 

epithelial cells (slightly Larger than pus cells), grouped in the 
form of a short cylinder and cemented together with a hyaline 
or mucoid matrix. The cells may be either whole or fragmented, 
clear, opaque, or granular. They are significant of an acute 
desquamative process, resulting from renal inflammation. 

Fatty Casts. — These casts may possess any of the charac- 
teristics of the preceding varieties, and present in addition free 
fat-globules scattered throughout the cast. They are considered 
as proof of fatty degeneration of the kidney. 

Blood Casts. — These casts are composed either of coagu- 
lated blood, in which innumerable corpuscles in various stages 
of disintegration are embedded, or they may represent a hyaline 
matrix in which appears a varying number of reel blood-cells. 
Some hyaline casts, which show a few blood-cells adherent upon 
their surfaces, may be simply hyaline or granular, to which the 
red cells have become attached in the bladder or after voiding. 
The presence of true blood casts indicates hematuria of renal 
origin. 

Bacterial Casts. — These resemble the granular variety, 
except that they are more closely and more uniformly granular. 
The bacteria may be so numerous that the cast is almost opaque. 
They denote the occurrence of an acute bacterial infection of 
the kidney, and are rarely found. 

Waxy Casts. — These are of rare occurrence, and are prob- 
ably simply a dense variety of the hyaline cast. 

Crystalline Casts.-— As the name implies, they are crys- 
talline in nature, being composed usually of uric acid, and more 
rarely of oxalates. They are very rarely encountered. 



CYLINDROIDS. 

These are narrow T , ribbon-like bands which usually present 
longitudinal striations, which may or may not extend through- 
out the entire length of the cylindroid. 

They are essentially hyaline in nature, and are probably 
formed from a hyaline basis. They are of renal origin, and are 
encountered in about 75 per cent, of cases which ordinarily come 
to the physician in the course of practice. 

Their significance is slight, though they are considered to 



222 THE URINE. 

indicate a degree of kidney irritation. In this connection it is 
of interest to note, that when found, they are many times accom- 
panied by a high specific gravity, and oxaluria, with or without 
the presence of indican. 

Microscopic Appearance. — Cylindroids appear as faint, 
ribbon-like bands, seen best by low illumination, and frequently 
exceeding in length the diameter of the field of the microscope 
when viewed through an objective of moderate power. Longi- 
tudinal striations may usually be detected, often running 
through only a portion of their length, the ends of which, after 
making one or more graceful curves, terminate in a gradual 
taper. 

Cylindruria with Albuminuria. — This condition may occur 
as a transitory phenomenon after such violent exercises as bicycle 
racing, foot-ball or rowing; also in chronic constipation or 
after the ingestion of moderate or large amounts of alcohol, par- 
ticularly by those unused to it. Continued ingestion of the 
salicylates has been found to produce it. It is often present 
during attacks of intestinal indigestion associated with oxaluria. 



SPERMATURIA. 

The presence of semen in the urine will give positive reac- 
tions for albumin. It may be present in the urine after coitus, 
after nocturnal emissions, or in spermatorrhea. 

Microscopic examination will reveal the characteristic ele- 
ments which are motionless, but which resist decomposition for 
days. 

CHYLURIA. 

Under very occasional circumstances the urine may contain 
chyle, which gives it the appearance of milk, and forms on 
standing a cream-like layer at the top. It responds to all the 
tests for albumin. 

Microscopically, much fat is found in the form of innumer- 
able highly refractive globules, which are soluble in ether. 



INORGANIC SEDIMENT. 



223 



THE INORGANIC SEDIMENT. 

To obtain a specimen of inorganic sediment, it is preferable 
to centrifuge the freshly voided urine rather than to wait for 
the slower process of sedimentation. The use of the centrifuge 
precludes the possibility of the process of decomposition so 
altering the specimen that the original picture is destroyed. 




ABC 

Fig. 30.— Centrifuge Tubes. 

A, Correct form of graduated percentage tube (note the slender taper which facilitates 

reading of small per cent.). B, Improper form of graduated tube accurate 

reading of small percentages difficult. C. Plain centrifuge tube. 

Alteration in reaction will so change the character of the crys- 
talline deposit that it will be useless from a diagnostic stand- 
point. A decomposed alkaline urine will often present such a 
mass of phosphatic sediment that the less voluminous, but more 
important, elements are greatly obscured and may be over- 
looked. 



224 THE URINE. 

Preparation of the Slide. — The sediment should be taken 
from the bottom of the centrifuge tube (Fig. 30) by means of a 
small pipette with a long, slender tip. Not more than a drop or 
two should be allowed to enter the tube. This is particularly 
important when the amount of sediment is small. If the sedi- 
ment is large and dense it should be diluted with a drop or two of 
distilled water and a cover-glass placed upon it. Much time will 
be saved by first examining the preparation with the % or y± 
objective, which is convenient for locating an interesting part 
of the slide, upon which the high-power objective may be focused 
for more careful study. 

Crystalline Deposits. (See Plates IV, V and VI.) 

Acid Group. — Uric Acid (Plates IV and VI) : These crys- 
tals are yellow, reddish-brown or brown in color. The most 
characteristic forms are rhombic prisms or lozenge-shaped crys- 
tals (Plate IV, Figs, a and b). These occur singly, but more 
often they are united in irregular masses. (Other more rare 
forms are shown in Plate IV, Figs, c, d, e and f; Plate VI, a 
and b.) 

Urates. — The urates, chiefly the urate of sodium and the 
urate of potassium, if they do not appear as an amorphous de- 
posit, appear as crystals in the forms of needles or dumb-bells, 
of reddish-brown color, and also in globular masses which are 
dark-brown and almost opaque, with or without projecting 
spines. 

Oxalates. — The usual form of calcium oxalate in the urine 
is a perfect octahedron without color. More rarely they appear 
in the conventional hour-glass form (Plate V, a and b). This 
form is somewhat similar to the urate from which it may be 
distinguished by the total absence of color in the oxalates. 

Carbonates. — These are rare, but when present evolve bub- 
bles of gas when treated with hydrochloric acid under the micro- 
scope. 

Sulphates. — This is a rare form of deposit which, when 
present, appears as fine, feathery crystals. Frequently a number 
of crystals radiate from a common center. 

Alkaline Group. — Phosphates: These may occur as a 
semi-opaque amorphous deposit without color. More commonly 
they appear as the characteristic coffin-lid crystals. A less com- 



PLATE IV. 




a and b. Calcium Oxalate Crystals. c, d, e and f. Phosphates. 



PLATE V. 




a and b. Uncommon Forms of Uric Acid Crystals. c. Cholesterin. 
d. Cystin. e. Tyrosin. f. Leucin. 



PLATE VI. 




a and b. Usual Forms of Uric Acid Crystals. c, d, e and f. Less 
Common Uric Acid Crystals. 



URINARY CONCRETIONS. 225 

mon form of crystalline phosphatic deposit (Plate V, c, d, e 
and /') appeals as line, branching, feathery, crystals, which have 
been likened to the needles and branches of the pine tree. 

Ammonium Urate (see Fig. 23). — These are characteristic 
of the uric acid and urate group in that they are yellow or 
brownish in color. In alkaline urine the urates appear as fine, 
feathery spheres of varying size, resembling to some extent 
chestnut burrs. 

Cli&lesterin. — This is a rare form of deposit which appears 
in the form of irregular, flat platelets whose sides follow the 
characteristic lines of a parallelogram, the angles of which are 
often irregular. Not infrequently the platelets occur in over- 
lapping groups. (Plate VI, c.) 

Cystin. — This is a rare deposit. When present it appears 
as irregular transparent plates of varying size often in over- 
lapping groups. (Plate VI, d.) 



METHOD TO DETERMINE ROUGHLY THE NATURE OF 
THE COMMON UNORGANIZED URINARY DEPOSIT. 

1. Warm the deposit and some urine. If it dissolves, the 
deposit is composed of urates. If it is unaffected by heat, the 
deposit is either phosphates, uric acid, or calcium oxalate. 

2. Warm a fresh portion of urine with acetic acid. If the 
sediment dissolves, it is composed of phosphates; if it does not 
dissolve, it is uric acid or calcium oxalate. 

3. Add to this undissolved portion some HC1 and heat 
again. If the sediment now dissolves, it is calcium oxalate; if 
it does not dissolve, it is composed of uric acid. 

URINARY CONCRETIONS. 

General Considerations. — About 75 per cent, of all urinary 
concretions are either uric acid or urates. Next in frequency 
are found the calcium oxalate or mulberry concretions. More 
rare primary concretions may consist of blood, cystin, xanthin, 
calcium phosphate, or calcium carbonate. 

Secondarily, any one of these formations may become cov- 
ered with a whitish layer of mixed phosphates. These are pre- 

15 



226 THE URINE. 

cipitated upon the original concretion by ammoniacal fermenta- 
tion, which occurs in the bladder, secondary to the presence of 
the calculus. 

Analysis of the Concretion. — 1. Burn a portion upon a 
piece of platinum foil in a Bunsen flame or blow-pipe. 

A. If it chars greatly and leaves little ash, it may be uric- 
acid, urates, cystin, xanthin or blood. 

1. If it gives the murexid test, it is uric acid or 

urates. 

2. If it dissolves in boiling water, it is urates. 

3. If it does not dissolve in boiling water, it is 

uric acid. 

4. Cystin and xanthin are very rare forms of con- 
cretions which require special tests for their 
detection. 

B. If it chars very slightly and leaves considerable ash it 
may be phosphates, oxalates or calcium carbonate. 

1. Treat a fresh portion with dilute HC1. 

(a) If it dissolves with effervescence, carbo- 

nates are present. 

(b) If it is soluble without effervescence. 

phosphates or oxalates are present. 
(See C, 3, below.) 

2. Treat a portion with dilute acetic acid. 

(a) If it dissolves with the aid of heat, it is 
phosphatic. 
C. — 1. If it fuses to a bead on platinum foil, it is urates. 

2. If it does not fuse, it is calcium phosphate. 

3. If it is insoluble in either of the above acids, it is calcium 
oxalate. 

THE DIAZO REACTION. 

The Reaction. — The ammoniacal solution of the anhydride 
of para-diazo-benzin-sulphonic acid has the property of devel- 
oping a salmon pink or red color reaction with certain patho- 
logic urines. Xormal urine never gives this reaction. 

The Test (for preparations of solutions see Appendix). — 
Take 5 cubic centimeters of solution "&" and add three drops 
of solution "B " and mix together in a clean test-tube. To this 



DIAZO REACTION. 227 

add 5 cubic centimeters of urine and again mix. Xow allow a 
few drops of ammonia water to flow down the side of the tube. 
As it conies in contact with the mixture in the tube a pinkish 
or red color indicates a positive reaction. Upon shaking the 
tube the entire mixture becomes colored, and the color is also 
imparted to the foam. This reaction does not always appear 
immediately, so a few minutes should be allowed for its devel- 
opment before reporting a negative reaction. 

Significance of the Reaction. — It is not yet definitely known 
what particular substance or substances in the urine yield this 
reaction. It may occur in the presence of several aldehydes, 
ketones, and phenols. 

A positive reaction is usually obtained with the urine of 
typhoid fever patients after the fourth day of the disease. Its 
continued absence, however, does not preclude the possibility of 
this disease. It is also positive in some cases of measles, pul- 
monary tuberculosis, etc. 

EXAMINATION OF THE URINE FOR SUBSTANCES INTRO- 
DUCED INTO THE BODY FROM WITHOUT 
(Drugs and Poisons). 

Detection of Lead. — A bright strip of magnesium, free from 
lead, is placed in the urine and left there for some time; the 
deposit which forms is dissolved in nitric acid, and then tested 
according to the methods of inorganic analysis. 7 

If the urine contains but a small amount of lead this method 
will not give the desired result ; then a larger quantity of urine 
must be used, the organic substances destroyed with HC1 and 
potassium chlorate, and the lead sought for in the evaporated 
residue. 

Detection of Mercury. — To 500 to 1000 cubic centimeters of 
urine, add 2 to 4 cubic centimeters of HC1; then digest at 60° 
to 80° C, for five or ten minutes in a flask with a few bright 
>trips of brass or copper. The metal is then washed with 
water, then with alcohol, and finally with ether, and dropped 
into an ignition tube (glass tube of high-fusing point). This is 
then brought to a red heat, care being exercised to keep the 



7 See Marshall's Medicus, J. B Lippincott Co. 



228 THE URINE. 

upper end of the tube cool. The mercury which has become 
amalgamated with the metal becomes volatilized, and is rede- 
posited in the upper cold end of the tube, where it is seen as a 
bright mirror, or, if a small amount, the individual globules of 
mercury may be seen by a hand-lens or the low power of the 
microscope. 

Detection of Iodine. — Iodine occurs in the urine as potas- 
sium iodide after the internal or external application of iodine 
or one of its combinations. It may easily be demonstrated as 
follows: A few cubic centimeters of urine are boiled with a 
piece of starch of about the size of a pea until the latter is dis- 
solved. After cooling, the fluid is carefully overlaid with con- 
centrated nitric acid. If iodine is present a blue-violet ring, 
which gradually disappears, is formed at the line of junction of 
the two fluids. A second method is to add to the urine a few 
drops of crude nitric acid and a less number of drops of chloro- 
form, and then shake gently. If iodine is present the 
chloroform, which sinks to the bottom of the tube, will be 
colored rose-red or violet. Both the above tests are very 
delicate, but if the urine contains only a small trace of iodine, 
the chloroform-test is not very conclusive, since the nitric acid 
may set free indol and skatol pigments, as well as uroresein, 
any of which will color the chloroform in a similar manner. 
But in this case the urine itself appears more deeply colored 
than the chloroform. 

Detection of Bromine. — The test for bromine is performed 
in exactly the same manner as iodine-test, except that a few 
drops of a calcium chlorid solution and hydrochloric acid are 
used to liberate the bromine. If bromine is present the chloro- 
form will be colored yellowish-brown. This test, though far less 
delicate than the iodine-test, is sufficiently accurate to recognize 
the therapeutic ingestion of large doses of bromine salts, and is 
chiefly of interest in verifying the diagnosis of suspected bro- 
mism. 

This method may be uncertain because of the discoloration 
of the chloroform by urinary derivatives. To prevent this source 
of error 16 cubic centimeters of urine, to which have been added 
2 cubic centimeters of caustic potash and 2 cubic centimeters 
of potassium nitrate, are evaporated and incinerated, the ash 



DRUG REACTIONS. 229 

dissolved in water, and the resulting solution tested for the 
presence of bromine, as above. 

Detection of Salicylic Acid. — Dilute ferric clilorid is added 
to the urine drop by drop. If the latter becomes a more or less 
intense violet, the reaction is positive. Salicylic acid and its 
salts, in which latter form the administered salicylic acid partly 
appears in the urine, both give this reaction. 

Detection of Phenol. — Phenol appears in the urine largely 
as phenol sulphate. Ferric chlorid will produce a violet-blue 
color in the distillate from phenol urine, to which 5 per cent, 
sulphuric acid has been added. Phenol urine turns dark or black 
on exposure to the air, when sufficient time is allowed for oxida- 
tion to occur. 

Detection of Antipyrin. — The urine appears dark and is 
dichroic, i.e., in reflected light, greenish; by transmitted light, 
reddish. A permanent brown-red color gradually appears upon 
the addition of ferric chlorid solution. 

Detection of Phenacetin (acetphenetedin). — The urine is 
dark yellow and turns reddish-brown on the addition of ferric 
chlorid solution. The color gradually becomes black after pro- 
longed standing. 

Detection of Antifebrin. — The urine is extracted with chlo- 
roform, and to the extract mercuric nitrate is added. The mix- 
ture is then heated, and if a green color is produced antifebrin 
is present. 8 

Detection of Pyramidon. — The urine is frequently clear, 
purplish-red in color, and deposits a sediment composed of 
small, red needles. If the urine is mixed with an equal volume 
of a 2-per-cent. solution of ferric chlorid, a dark-brown, ame- 
thyst color is produced. 

Detection of Tannin. — Tannin is eliminated in the urine 
partly as gallic acid. Urine which contains tannic and gallic 
acids turns a deep blue-black upon the addition of ferric chlo- 
rid solution. 

Detection of Balsam of Copaiba and Sandalwood Oil. — After 
the administration of copaiba the urine will reduce cupric oxid 
(Trommer's test), but not bismuth ( Nylander's test). If HC1 



8 Yvon. Jour, de Pharm. et de Chemie. No. 1, 1887. 



230 THE URINE. 

is added to the urine drop by drop, a precipitate of resinous 
acids appears with a reddish or violet coloration. Also after the 
administration of sandalwood oil the urine has reducing prop- 
erties, and exhibits a precipitate of resinous acid when HC1 is 
added, but it will be of a reddish-brown color. 

Detection of Santonin. — Santonin urine is of a saffron-yel- 
low or greenish color. The addition of sodium hydrate will 
turn it a rose-reel. If this rose pigment is shaken with amylic 
alcohol, it will be immediately taken up by the latter, giving it 
an intense and beautiful color, while at the same time the urine 
is decolorized. 



XII. 
THE CEREBROSPINAL FLUID. 



To Obtain the Specimen. 1 — It is very important when with- 
drawing the cerebrospinal fluid to see that too much is not ab- 
stracted at one time. Death has resulted from too rapid and 
too great removal of fluid. This risk need hardly be considered 
in obtaining fluid for purposes of examination, as not more than 
8 to 10 cubic centimeters are required. Untoward symptoms 
are less likely to follow spinal puncture if the patient is kept 
in bed for a few hours succeeding the procedure. 

The control of the needle is important. It is quite possible 
with an 8-centimeter needle to thrust into the peritoneal cavity 
or between the bodies of the vertebra, and by this latter path to 
pierce even the vena cava. Aspiration to start the flow is not 
permissible, as with proper precautions and careful technic a 
dry tap is very rare. 

The Pressure. — For exact pressure-determinations elaborate 
apparatus is not required. For ordinary bedside work the ap- 
paratus about to be described is sufficient : As soon as the sub- 
arachnoid space' has been entered, as shown by the appearance 
of fluid in the needle, a thick-walled glass tube of fine bore 
(about l 1 /^ millimeter diameter) is joined to the needle by a 
short rubber connection, and the fluid allowed to find its level 
in this tube held vertically, and the pressure measured in terms 
of millimeter according to the height to which the fluid rises. 
The capillary error, variations due the differing specific gravity 
in the fluid, and the change due to loss of fluid from the spinal 
canal, may be neglected for all practical purposes. Under ordi- 
nary bedside conditions, with this apparatus, the normal varia- 
tions in cerebrospinal pressure are between 50 and 300 milli- 



1 Abstract of article by Francis Peyton Rous, M.D., in International Clinics, Vol 
ii, Seventeenth Series. 

(231) 



232 THE CEREBROSPINAL FLUID. 

meters of the fluid itself. By careful experimentation and elabo- 
rate apparatus, the normal variations have been found to lie 
between 120 and 180 millimeters of distilled water. 

Physiologic Modifications. — Crying or coughing during the 
examination, will cause a rise in pressure of 50 millimeters 
or more. Posture also influences pressure, as is shown if the 
patient (who should be reclining horizontally on one side) raises 
his head. The column further shows fluctuations synchronous 
with the pulse and respiration. Finally the absolute pressure 
will be higher than normal if the patient exerts any muscular 
force during the examination. Muscular exertion should be 
eliminated as much as possible during the reading. 

Pathologic Modifications. — Marked increase in pressure is 
the rule in hydrocephalus, in brain tumor, and in meningitis of 
bacterial origin, especially when the infection is acute. Increase 
in pressure often accompanies uremic coma. A column of 600 
to 700 millimeters of fluid is not uncommon in any of these con- 
ditions. Low pressure is observed in infants in conditions giving 
low-blood pressure (see section on "Blood-pressure"), and when 
an error in technic has occurred. 

Collection of the Specimen. — When the pressure has been 
determined, disconnect the manometer and collect a few cubic 
centimeters in a sterile capillary tube as the fluid emerges from 
the needle. This specimen should be preserved under aseptic 
precautions for bacteriologic examination; from 4 to 6 cubic 
centimeters more should be allowed to run directly into a clean 
centrifuge tube for cytologic examination. 

Determination of the Cell-Content. — In the normal cerebro- 
spinal fluid there are from one to seven white cells per cubic 
millimeter. Probably they are never entirely absent. When the 
meninges are sound none but lymphocytes occur ; these are often 
much degenerated. The fluid is normally clear, and even if it 
contains several hundred cells per cubic millimeter, it may re- 
main clear macroscopically. 

The cells may be counted with the Thoma-Zeiss hemo- 
cytometer, using the "red" pipette for measuring. Since there 
is usually some hemorrhage accompanying the puncture, it is 
necessary to have previously counted the number of red and 
white cells in the patient's blood to determine the ratio of these 



DIFFERENTIAL COUNT. 233 

cells in the circulation, so that alter counting the red and white 
cells in the cerebro-spinal fluid it will be possible to determine 
how many white cells are adventitious (due to hemorrhage). 
To determine the cell-content of the cerebrospinal fluid it is only 
necessary to place a drop of the freshly drawn fluid upon the 
depression in the chamber and apply the cover-glass; to count 
the red cells one filling will be all that is necessary. For the 
white cells, if the cells are scant, five or ten complete fields 
must be counted and an average obtained. 

Example. — Suppose in the specimen we find 80,000 red 
cells and 102 white cells per cubic millimeter, while in the blood 
itself there are 4,000,000 reds and 5000 whites, or a proportion 
of 800 to 1. Calculating on this ratio of the 102 white cells 
present per cubic millimeter of the cerebrospinal fluid, 100 are 
from the contamination blood. Thus, but two white cells per 
cubic millimeter were present in the fluid before contamination 
by hemorrhage. 

Pathologic Variations in the White Cells. — The cell-content 
of the cerebrospinal fluid is an extremely sensitive index of the 
state of the meninges. 

Acute Mexixgitis. — Purulent fluids may contain from 
4000 to 40,000 white cells per cubic millimeter. 

Tubercular mexix^gitis will average between 200 and 300 
cells per cubic millimeter, although it may be as high as 20,000. 

Syphilitic mex^ix^gitis, tabes, paresis, usually give more 
than 10 and less than 100 cells per cubic millimeter. 

Caution. — It must be borne in mind that the occurrence of 
a slight increase in the number of white cells may be the result 
of a previous puncture. This question should be ascertained 
before arriving at final conclusions in any case. 

It must be remembered, further, that in cerebrospinal fluid 
kept at room-temperature for a short time, the cells rapidly de- 
generate so that they are often unrecognizable after a lapse of 
a few hours, even if they have not entirely disappeared. 

Differential Count. — In general, should the pressure of the 
fluid be between normal limits, and the cells less than 10 per 
cubic millimeter, it is unnecessary to make a differential count. 
Under these circumstances lymphocytes alone are present. 
The differential count should be accomplished speedily after 



234 THE CEREBROSPINAL FLUID. 

removal of the fluid to prevent alteration due to decomposition. 
The fluid which has been allowed to fall into the centrifuge 
should be revolved rapidly for from ten to thirty minutes. This 
completed, the tube should be carefully removed from its metal 
sheath to avoid dissipating the cells. To transfer the sediment 
to a slide or cover-glass the fluid must be drained away by 
slowly and steadily inverting the tube. While the tube is still 
inverted the slight portion of fluid remaining in the end of the 
tube is taken up on a capillary pipette and blown out on to the 
previously cleaned glass. This is air-dried, when it is ready for 
fixing or staining (for methods of staining for differential count 
see section on "The Blood"). 

Lymphocytes alone (and perhaps occasionally a large, flat 
endothelial cell) are normal to the fluid. In the differential 
count from the stained specimen, it is necessary to count and 
classify the polymorphonuclears, the mononuclears, and endo- 
thelial cells. Mast cells, eosinophiles, tumor or nerve cells, have 
rarely been detected, and must still be considered exceptional, if 
not doubtful. The relationship of the polymorphonuclears and 
the lymphocytes are alone of importance in our present state of 
knowledge. 

Polymorphonuclears are indicative of an acute process, 
while the mononuclears speak for chronicity. The mononuclears 
(lymphocytes of blood) appear to be particularly numerous in 
tubercular meningitis and cerebrospinal syphilis. 

Determination of Proteid Content. — The fluid remaining 
after the bacterial examination, and that supernatant after cen- 
trifugation, may be devoted to the proteid determination. An 
albumin and a globulin have been found in the cerebrospinal 
fluid, the relative importance of each has not yet been deter- 
mined, so for clinical purposes their combined content alone is 
determined. The acetic-acid boiling-test may be applied here, 
provided the examiner is familiar with the normal amount of 
proteid as represented by the white cloud formed in the upper 
half of the tube. For a rough quantitative determination a very 
narrow test-tube may be marked off in lengths to correspond to 
those of the Esbach tube. The Esbach reagent is used as in the 
similar test for albumin in urine. This test will serve to show 
the proteid variations from time to time in a given case. 



INTERPRETATION OF FINDINGS. 235 

There is an increase in the proteid content of the cerebro- 
spinal fluid in all inflammations of the meninges. Proteid 

increase ami cell increase usually go hand in hand, but this 
relationship is not constant. 

Interpretation of Findings. — Having ascertained the pres- 
sure, the cell-content and cell-character, and the amount of pro- 
teid in any given case, one is in a position to state much. 

1st. As to the presence or absence of meningeal infection, if 
the fluid be clear and colorless macroseopically, the cells less than 
S per cubic millimeter, and the proteid content low with the 
pressure between 50 and 300 millimeters of the fluid itself, this, ' 
as far as is at present known, may be considered conclusive evi- 
dence against an active pathologic change in the meninges. 

*2d. A red-yellow or brown fluid under increased pressure 
(above 300 millimeters) showing erythrocytes, and increased 
proteid content, is evidence pointing to hemorrhage into the 
subarachnoid space. 

3d. When tabes, paresis or cerebrospinal syphilis is sus- 
pected, a specimen showing moderate cell increase (10 to 200 
cells per cubic millimeter), mononuclear in character, with or 
without proteid increase, and the pressure not above 300, speaks 
for a chronic process. 

4th. When meningitis is in cmestion a definite increase in 
cells (over 100), specially of the polymorphonuclear variety, 
increase in proteid, and pressure greater than 300 millimeters, 
is strongly in favor of a diagnosis of acute meningitis, even in 
the absence of macroscopic change in the fluid and a negative 
bacteriologic report. 

5th. Cases of brain tumor and of hydrocephalus usually 
show an increase in pressure, with a normal cell-content. 

Bacteriologic. — For staining method see section on "Bac- 
teriologic Methods." 



XIII. 
THE BODY FLUIDS. 



THE PERITONEAL FLUID. 

Characteristics of Normal Fluid. — In conditions of health 
there is just sufficient fluid to thoroughly lubricate the interior 
of the abdominal cavity. This fluid is clear, of a pale, straw 
color, having a specific gravity of 1005 to 1015; is slightly 
albuminous and, under the microscope, a drop of freshly-drawn 
fluid placed between a clean slide and cover-glass, reveals very 
few, if any, formed elements. 

Pathologic Exudate. — Inflammatory ascitic fluid is 
straw or lemon-yellow in color, and usually somewhat cloudy, 
depending upon the number of cells contained in it. Its specific 
gravity varies between 1012 and 1026, or even higher. The fluid 
frequently coagulates spontaneously. It shows a variable amount 
of albumin by boiling, and sugar, bile-pigments, urea, uric acid, 
and cholesterin may be demonstrated by appropriate means (see 
section on "Urine"). More rarely xanthin, creatinin, and allan- 
toin may be found. 

Hemorrhagic effusion occurs in cancer and tuberculosis 
of the abdominal cavity, and occasionally in cirrhosis of the 
liver. 

A chylous or milky fluid is occasionally met. This is 
readily distinguished by examination of fresh fluid under 
the microscope, when the characteristic fat globules will be 
found. 

Non-Inflammatory Transudate. — In Bright's disease the 
ascitic fluid is usually a pale clear serum of low specific gravity, 
showing a minimum of cells. 

In cirrhosis of the liver the color is usually darker and bile- 
pigment can be demonstrated. 

Differentiation. — Ovarian Cysts : The fluid obtained 
from ovarian cysts has a specific gravity usually above 1030, and 
(236) 



PLEURAL FLUID. 237 

is of a dark-brown, grumous appearance. Ovarian fluids are fur- 
ther said to contain a Large number of compound granule cells 
which, from their frequent occurrence in this fluid, have been 
termed "ovarian cells." Microscopically these appear as Large 
oval or round cells, showing dense, irregular granulations often 
obscuring the nucleus of the cell. 



PLEURAL FLUID. 

General Considerations. — Physiologically there is not pres- 
ent enough fluid for analysis. Under pathologic conditions it 
may vary from one to four or more liters, and may be serous, 
sero-purulent, purulent or hemorrhagic. 

Non-Inflammatory Transudate. — In hydrothorax the spe- 
cific gravity, as a rule, is below 1015, the albumin content is 
small, and there is little tendency for the fluid to coagulate spon- 
taneously. The fluid is clear or pale, straw-colored, unless 
tinged with hemorrhage occurring during the puncture. The 
formed elements are very scarce and hard to demonstrate. 

Inflammatory Exudates. — Sero-Fibri^tous Pleurisy : The 
serous exudate is abundant, and flakes of fibrin are present in 
the fluid, appearing as fibrillated flocculi. The actual amount of 
fluid varies greatly. It is of a citron color, either clear or slightly 
turbid from the fibrin and formed elements present. In some 
cases the color is dark-brown, usually due to altered blood con- 
tained in it. Specific gravity below 1020. 

Microscopically, we find a variable number of leucocytes 
and red blood-cells, and some swollen endothelial cells and bac- 
teria. 

Purulext Pleurisy. — The specific gravity is usually above 
1020. The fluid has a heavy, sweetish odor, which, if the in- 
fection be due to a penetrating wound, may be fetid. The fluid 
is essentially pus, contains a very large amount of albumin which 
may coagulate spontaneously after removal. Appropriate tests 
may show cholesterin, uric acid, bile-pigment, and sugar. 

Method of Making Permanent Stained Preparation. — In 
fluids which are clear and show but few cells on microscopic 
examination of the fresh specimen, it becomes necessary to cen- 
trifuge 15 to 20 centimeters in order to concentrate these formed 



238 THE BODY FLUIDS. 

elements. After centrif ligation in the ordinary urine centrifuge 
for from two to three minutes, the supernatant liquid is slowly 
and steadily poured off, and the remaining residue taken up in 
a capillary pipette from which it is blown out upon clean cover- 
slip and allowed to dry spontaneously (without heating). It 
then may be stained by any of the Romanowski stains (see sec- 
tion on "Blood") or the eosin-methylene-blue sequence. 

Fluids which contain cells in macroscopic amount may be 
transfered to cover-glasses for staining without centrifugation. 

Microscopic Examination of Stained Preparations. — In 
hydrothorax the cells are few and mainly large, flat, of endothe- 
lial origin. 

In pneumo- and streptococcic pleurisy, there is a great pre- 
ponderance of polymorphonuclears. In tubercular inflamma- 
tions the lymphocyte is the predominating cell. 

Many efforts at classification and diagnosis have been made 
on a limited number of examinations with a view to arriving 
at some definite rules for cytodiagnosis of pleural effusions. 
Further systematic study along these lines is, however, neces- 
sary. 

The presence of red cells is usual in tuberculous processes, 
also in the presence of rupture of an abscess from an adjacent 
organ into the pleural cavity. Very frequently red blood-cells, 
in a pleural effusion, are due to the traumatism inflicted at the 
time of puncture. Bed cells which appear in the first few cen- 
timeters drawn, are most likely of this origin. 

The possibility of diagnosing cancer by microscopic exami- 
nation of the fluid is always present, through the finding of 
characteristic cell masses in the fluid. 

THE PERICARDIAL FLUID. 

This fluid normally is of a pale, lemon-yellow color, slightly 
viscid, cloudy from cell detritis; occasionally it may be clear. 
It contains a moderate amount of albumin and certain inorganic 
salts. The specific gravity ranges between 1015 and 1030. 

THE SYNOVIAL FLUID. 

This fluid is alkaline, thick, viscid, sticky, and of a yellow- 
ish color. Physiologically the joints contain just sufficient to 



HYDROCELE. 239 

completely lubricate them. Under pathologic conditions the 
fluid, in a large joint, may amount to many cubic centimeters. 

HYDROCELE FLUID. 

The fluid is usually clear, of a yellow or greenish tinge. 
The specific gravity varies between 101-i and 102(3. The fluid 
sometimes coagulates spontaneously. Some leukocytes are 
always present, and occasionally crystals of cholesterin. 



XIV. 

HUMAN MILK. 



General Considerations.— It must always be remembered, 
in the examination of milk, that it is no simple matter to obtain 
a truly representative sample. The fatty matters tend to sepa- 
rate, and there may be great variations between different por- 
tions of the same sample. This difficulty is particularly pro- 
nounced when human milk is examined, because of changes in 
the milk incident to mental and nervous disturbance. 

The Sample. — The sample should consist of a thorough 
mixture of portions taken at separate intervals, by means of the 
breast-pump. Wherever possible it is well to have the woman 
somewhat accustomed to the use of the pump before taking any 
milk for examination. In all cases the sample should be thor- 
oughly mixed before testing, by pouring rapidly from vessel to 
vessel. 

Physical Characteristics. — Woman's milk is bluish-white in 
color, of sweetish taste and characteristic odor. When freshly 
drawn it is alkaline or amphoteric, but never under healthy 
conditions is it acid. The specific gravity varies between 1026 
and 1036, the average being 1031 at 60° F. On the addition 
of acetic acid only a slight coagulum is seen, being in the form 
of small, fine flocculi, never in large masses as is the case with 
cows' milk. Besides the myriad fat-globules, may be seen large, 
flat epithelial cells from the milk ducts. 

Composition of Human Milk (after Holt). 

Average. .Normal variations. 

Fat 4.00 per cent. 3.00 to 5.00 per cent. 

Sugar 7.00 " 

Proteids 1.50 " 



Salts 0.20 

Water 87.30 

Total 100.00 

(240) 



6.00 to 7.00 

1.00 to 2.25 

0.18 to 0.25 

89.82 to 85.50 

100.00 to 100.00 



DETERMINATION OF THE FAT. 241 

The composition and food value of human milk are im- 
paired when the mother is unhealthy. It is affected injuriously 
when there exists undue emotional excitement. It will contain 
an appreciable amount of certain medicines ingested by the 
mother, which may affect the infant. 

Examination of the Milk. — The exact determination of the 
composition of human milk is only to be determined by a com- 
plete chemical analysis. There are, however, many variations 
from the normal which the physician may readily ascertain for 
himself by simple methods of examination. 

The Quantity. — This may be determined roughly by using 
the breast-pump, although this is not reliable for many reasons. 
With sufficiently sensitive scales the physician may, by weigh- 
ing the infant before and immediately after nursing, determine 
whether it is getting only one or two, or four or five ounces. The 
average daily quantity secreted by woman is one liter or two 
pints. 

The Eeaction. — Test by litmus paper. The reaction 
should be alkaline or amphoteric, but never acid when freshly 
drawn. 

A spontaneous change occurs after milk has stood for some 
time in a warm place; it coagulates or sours, and becomes acid. 
This is due to the change of milk-sugar (lactose) into lactic 
acid. This occurs through the agency of the bacterium lactis. 
The casein normally is held in solution by the alkaline phos- 
phates, the acid changes the reaction, and hence causes the 
casein to be precipitated. 

Specific Gravity. — This may be taken by a small hydro- 
meter which is graduated from 1010 to 10-10. The specific 
gravity is lowered by fat, but increased by the other solids. 

Microscopically, milk is found to be composed of minute 
brilliant, oil globules encased in a thin envelope of casein. 
Immediately after delivery the milk is relatively poor in casein, 
but rich in fatty matter, which exists in considerable amount in 
the form of colostrum masses. The microscope also reveals the 
presence of colostrum corpuscles, blood, pus, epithelium, and 
granular masses. Colostrum corpuscles are abnormal after the 
twelfth day; blood and pus are always abnormal. 

Determination of the Fat. — The simplest method is by 



242 HUMAN MILK. 

means of the cream gauge. This consists of a graduated test- 
tube with a foot, and a ground-glass neck and stopper. The 
tube is filled to the zero mark with freshly-drawn milk and 
allowed to stand at room-temperature for twenty-four hours, 
when the percentage of cream is read off directly from the grad- 
uated scale. The relation of cream to fat is approximately five 
to three. Thus 5 per cent, cream equals 3 per cent, fat, etc. 

Centrifuge Method. — The use of a specially graduated 
centrifuge tube is more accurate. These tubes may be used in 
the ordinary centrifuge for urine. Only 6 cubic centimeters of 
milk are required for this test. If carefully conducted the test 
is nearly as accurate as the chemical analysis. It gives results 
accurate to within one-fifth of 1 per cent. 

In the usual apparatus two pipettes are supplied with the 
centrifuge tubes, one of 5 cubic centimeters capacity, marked 
milk, the other holding 1 cubic centimeter up to a mark, for 
introducing the alcoholic solution. 

To determine the fat by this method, 5 cubic centimeters 
of the sample is introduced into the tube by means of the 
pipette marked "milk," 1 cubic centimeter of the alcoholic solu- 
tion (solution-A) is added, and the tube well shaken. Then, by 
means of any large pipette, solution-B is added little by little 
until the tube is filled to the zero mark. It is then placed in 
the centrifuge and rotated very rapidly for four or five minutes. 
This will bring the fat to the top in a clear yellowish layer 
which can be read off in direct percentage by the scale on the 
neck of the tube. 

A few drops of water may be added, if necessary, to correct 
the level of the top of the fluid. If the milk should be richer 
than 5 per cent, for it will be necessary to dilute the sample by an 
equal quantity of water, proceed with the test as above, and 
finally multiply the result by two. 

Solution-A consists of: — 

Amylic alcohol 37 parts by volume. 

Methyl alcohol 13 parts by volume. 

Hydrochloric acid 50 parts by volume. 

This solution may be kept for a short while ; if it turns dark 
it is worthless. Solution-B consists of sulphuric acid, specific 
sravitv 1832. 






ESTIMATION OF THE PROTEIDS. 



243 



Determination of the Sugar. — The percentage of sugar is 

nearly constant, so it may be ignored in the usual clinical in- 
vestigation. 

Estimation of the Proteids. — There is no simple method 
of determining the percentage of proteids. If we regard the 
sugar and salts as constant, or so nearly so as not to effect the 
specific gravity, we may form an approximate idea of the pro- 
teids from a knowledge of the specific gravity and the percentage 
of fat. We may thus determine whether they are greatly in ex- 
cess or very scanty. The specific gravity then will vary with 
the proportion of proteids, directly, and inversely with the pro- 
portion of fat, i.e., high proteids, high specific gravity, low fat, 
low specific gravity. The application of this principle will be 
seen by reference to the accompanying table. 1 



Composition of Woman's Milk. 

Proteids 
Specific gravity at 70° F. Cream 24 hrs. Calculated. 

Average 1031 7 % 15 %- 

Normal variation 1028-1029 8% to 12% Normal (rich 

milk). 

Normal variation 1032 5 to 6% Normal (fair 

milk). 
Abnormal variation. Low (below 1028) High (above Normal or 

10%) slightly 

below. 
Abnormal variation. Low (below 1028) Low (below Very low. 

5 % ) very poor 

milk. 
Abnormal variation. High (above 1032) High Very high, 

very rich 
milk. 
Abnormal variation. High (above 1032) Low Normal or 

nearly so. 

1 From Holt's "Pediatrics.' 



XV. 

BACTERIOLOGIC METHODS. 



By a gradual process of evolution and development the 
field of clinical medicine has so enlarged its borders that to-day 
the laboratory worker no longer confines his investigations to 
examination of the blood, urine, and sputum, etc., but must pos- 
sess a working knowledge of bacteriology and be familiar with 
bacteriologic technic to the extent that he may be able to obtain, 
differentiate, and recognize the commoner pathogenic organ- 
isms. This section will confine itself to a brief outline and dis- 
cussion of the essentials of laboratory technic, referring the 
worker, who would delve further into this fascinating field, to 
works devoted to bacteriology. 

The Tubercle Bacillus. — This is the most important organ- 
ism, from a clinical standpoint, at least, which is encountered 
in the field of clinical medicine. 

The tubercle bacillus presents great difficulties in the way 
of artificial cultivation, and the results of such procedures are so 
unreliable that they are rarely depended upon for purposes of 
identification, the diagnosis usually being made upon the greater 
acid-fast properties of this organism as compared with all 
others; the final corroboration of the diagnosis being made by 
animal inoculation, for which purpose the guinea-pig, owing to 
its great and uniform susceptibility, is commonly employed. 

The tubercle bacillus is, in the strict sense of the word, a 
parasite which finds conditions entirely favorable to its growth 
and reproduction only in the body. On artificial media the 
bacilli, even when transferred directly from the human body, 
grows only imperfectly or not at all. 

For artificial cultivation of the tubercle bacillus, the best 
results are obtained by the use of some blood-serum medium 
(see page 267). 

The finding of tubercle bacilli in the sputum is positive 
(244) 



STAINING METHODS. 245 

evidence of pulmonary, bronchial or laryngeal tuberculosis. On 
the contrary, their absence, after careful search, even of a num- 
ber of preparations, cannot be considered absolute negative evi- 
dence. 

If, after careful search of suspicious particles taken from 
the sputum of a suspect, whose clinical history and symptoms are 
strongly suggestive of the disease, the organism is not found, 
the following devices may be resorted to : — 

A. The whole amount of a twenty-four hours' specimen of 
sputum, to which no antiseptic solution has been added, should 
be placed in a porcelain dish or glass beaker, and stirred with a 
glass rod until quite thin and diffluent. This should then be 
stood aside for a few hours to settle, when the lowest portion 
of the fluid is taken up in a pipette and transferred to a centri- 
fuge-tube, where it is rotated for from fifteen minutes to half 
an hour. From the bottom of the centrifuge-tube a small 
amount of the sediment is transferred to a cover-glass or slide 
for fixing and staining. 

B. A considerable quantity of sputum is mixed with an 
equal quantity of water, and a few drops of a 10-per-cent. solu- 
tion of sodium hydrate is added. This mixture is then heated 
until homogeneous, when it is cooled, sedimented, and centri- 
fuged, and examined as outlined above. 

Microscopic Appearance. — The tubercle bacillus is a delicate 
rod usually appearing in stained specimens with a beaded in- 
ternal structure. It may be straight, but is usually slightly 
curved, in its long axis. Its length is very variable, some being 
short, others quite long, though never as long threads. Its aver- 
age length varies between 2 and 5 micro-millimeters, and it is 
usually very slender. 

In sputum the organisms may occur singly or in groups of 
from three to half a dozen or more. 

Staining Peculiarities. — The recognition of the tubercle ba- 
cillus depends upon a special method by which they alone are 
stained. Unstained they cannot be differentiated from the 
other organisms which may be present. The ordinary methods 
employed for staining bacteria are not suitable, so that special 
technic has been devised and is now regularly employed to render 
their recognition less difficult. 



246 BACTERIOLOGIC METHODS. 

In these methods advantage is taken of the fact that cer- 
tain substances increase the activity of staining by aniline dyes. 
With the tubercle bacillus this is accomplished with carbolic 
acid. Another important point is that these organisms, when 
once stained, give up their color only with great difficulty, so that 
agents, which will decolorize all other bacteria in the course of 
a few minutes, will have no appreciable effect upon the tubercle 
bacillus. It is upon these two peculiarities that we rely in dif- 
ferentiating this organism. 

Differential Diagnosis. — While the peculiar micro-chemical 
reaction toward staining reagents is unually considered to be 
unique with the tubercle bacillus, it should be remembered that 
at least three other species of bacteria, when similarly treated, 
react in the same way. This fact is particularly important in 
connection with the microscopic examination of urine and patho- 
logic secretions from the genito-urinary tract, and from the rec- 
tum. Here is commonly encountered the smegma bacillus which 
is the next most important member of the group of acid-fast 
organisms. Acid-fast bacilli have been found in the sputum 
and about the teeth and tonsils in a case of non-tubercular dis- 
ease of the lung. 1 

While these other organisms have the same acid-fast prop- 
erty as has the tubercle bacillus, they appear so rarely that seri- 
ous mistakes are not likely to occur. The method of Pappen- 
heim (see page 247) may be relied upon to overcome this diffi- 
culty. This method colors the tubercle bacillus red and the 
smegma bacillus blue. 

Staining Methods. — Ziehl-Nielsen : Place a few drops 
of the carbol-fuchsin solution (for preparation of stain see 
Appendix) upon the fixed cover-glass preparation, and hold 
over a low Bunsen flame until steam begins to rise. Do not boil. 
Continue the steaming process for from three to five minutes, 
pour off excess of stain, and wash in water. It is important to 
prevent the staining reagent from reaching the under surface 
of the cover-glass. If this is permitted the pigment will become 
dry and burned into the glass, when it will successfully resist all 
efforts at complete decolorization. Decolorize with acid-alcohol 



Pappenheim: Berlin, klin. Woch.. No. 37, p. 809. 



STAINING METHODS. 247 

solution (see Appendix) or with 25 per cent, sulphuric acid. De- 
colorizatioD should be continued until the red color has entirely 

disappeared from the specimen; if this process is not thoroughly 
done the finished slide, so far as its diagnostic value is con- 
cerned, will be worthless. The specimen is then washed in water, 
and counterstained for from one to three minutes with a 
1-per-cent. aqueous solution of methylene-blue, after which 
it is washed, dried, and mounted for microscopic examination. 
By this method the tubercle will appear as the characteristic 
red rods upon a blue background. A field which reveals other 
red objects has been insufficiently decolorized and should be dis- 
carded for a more carefully prepared specimen. 

Gabbett's. — Flood the dried and fixed specimen with car- 
hol-fuchsin and steam for three minutes; then pass directly to 
the acid methylene-blue stain (see Appendix) for one minute. 
Finally wash, blot, dry in the air, and mount. This field, if 
properly prepared, should have the same appearance as that- 
prepared by the Ziehl-Nielsen method. 

Pappexheim recommends the following technic : 1. Stain 
in carbol-fuchsin by steaming near the boiling point for three 
or four minutes. 2. Pour off the excess of carbol-fuchsin and 
treat without washing with Pappenhemr's solution (see Appen- 
dix), pouring it slowly three or four times over the preparation, 
and allow it to drain off. 3. Wash in water, dry, and mount. 
Duration of entire procedure from three to five minutes. 

Czaplewsky's Method. 2 — This method employs the fol- 
lowing solutions: (a) One gram of fuchsin is dissolved in 5 
cubic centimeters of liquid carbolic acid in a dish; 50 cubic 
centimeters of glycerin are then added with constant stirring, 
and finally dilute with water to 100 cubic centimeters. The 
solution is said to keep extremely well, and does not need 
to be filtered, (b) Ebner^s decolorizing solution (see Appendix). 

Method. — 1. Stain, with the aid of heat, for three or four 
minutes. 2. Decolorize by treating the specimen alternately 
with Ebner's fluid and distilled water.. 3. Counter-stain with 
methylene-blue for one minute. 4. Wash, dry and mount in 
Canada balsam. The chief advantage of this method is said to 



2 Hyg. Rundschau, No. 21. 1896. 



248 BACTERIOLOGIC METHODS. 

be that all acid-resisting bacilli but the tubercle bacillus, are 
decolorized, and therefore cannot be confused with the bacillus 
in question. 

DIPLOCOCCUS PNEUMONII. 

The presence of Frankel's diplococcus in the sputum of 
patients suffering with croupous pneumonia, is fairly constant, 
so that its demonstration is of considerable diagnostic impor- 
tance. They appear as elongated lanceolate cocci, usually ar- 
ranged in pairs with their bases approximated. They are sur- 
rounded with a faintly staining capsule which, in dry prepara- 
tions, does not usually take the stain at all, although the ordi- 
nary method employing methylene-blue has occasionally, in the 
author's experience, demonstrated a faint but distinct capsule. 

The organism is supposed to be the cause of lobar pneu- 
monia, but must not be confounded with the other diplococci 
occurring in the sputum, more especially with Friedlander's 
bacillus. The latter also possesses a capsule, but has nothing 
whatever to do with the production of lobar pneumonia, though 
occasionally they may accidentally be present. Friedlander's 
organism, when highly magnified, will be found to be a short 
rod. Cultural characteristics will also serve to differentiate, 
as will also Gram's staining method, which decolorizes Fried- 
lander's and stains Frankel's organism. 

The following modification of Gram's method will be 
found satisfactory (for preparation of staining reagents see 
x^ppendix) : Hold the fixed and dried cover-glass preparation 
in the forceps and flood with carbol-gentian violet; allow this 
stain to act for from three to four minutes. Wash and trans- 
fer to the iodine-potassium-iodide solution for from one to two 
minutes. Xext wash in alcohol until the apparently dirtily- 
stained film is decolorized. Transfer to absolute alcohol, then to 
oil of cloves, and finally mount in balsam. By this method the 
organism appears as a dark-blue or violet diplococcus. Fried- 
lander's organism will remain unstained. 

A very useful method for differentiating Frankel's coccus 
is that devised by W. Wolf. By this method the dry preparation 
is first stained in aniline water saturated with fuchsin, and 
then placed for one or two minutes in a dilute watery solution 



BACILLUS OF INFLUENZA. 249 

of methylene-blue. The cocci will now be found stained blue, 
the capsule rose-red, and the body of the specimen purplish-red. 

Method for Staining Capsules. — Prepare the cover- 
glass smear in the usual way, then without drying flood with 
glacial acetic acid. At the expiration of one minute pour off 
the excess of acid, and without washing flood the specimen with 
aniline water gentian-violet (Koch-Ehrlich), which should be 
allowed to act for four minutes, w T hen the excess of stain is 
poured off and a fresh portion added, which is allowed to act 
for another two minutes. The cover-glass is now washed in two 
or three changes of normal saline solution (it may be found 
necessary to employ a saline solution of 1.5 to 2.0 per cent.), 
after which it is blotted, dried, and mounted in the usual way. 

By this method, the capsule will appear as a faintly tinted 
halo about the diplococcus. The absence of this cocci practically 
excludes the diagnosis of lobar pneumonia, although its demon- 
strated presence is by no means positive evidence in the other 
direction, because this organism has repeatedly been demon- 
strated in the sputum and mouth secretions of healthy indi- 
viduals. 

BACILLUS OF INFLUENZA OR PFEIFFER'S BACILLUS. 

A small slender bacillus occurring usually in very great 
numbers in the nasal secretion of fresh attacks of true influenza, 
but not found in the ordinary short attacks of prostration accom- 
panied by coryza, which is at present designated influenza or 
"grippe," largely for lack of a more definite diagnosis. Mor- 
phologically, it is a very small rod appearing frequently in pairs ; 
in parts of the secretion this organism may be found in what is 
practically a pure culture, occurring both within and without 
the leukocytes. Their length is usually from two to three times 
their width, they rarely form chains. The ends of the rods are 
rounded off, and if imperfectly stained (they stain with diffi- 
culty) the ends will be more deeply stained than the center, 
giving an appearance not unlike a diplococcus with just a sug- 
gestion of a capsule. According to recent authority this bacillus 
does not possess a capsule, the deceptive appearance being prob- 
ably a staining peculiarity of certain cells. The bacillus is non- 
motile, and can only be cultivated upon special media contain- 



250 BACTERIOLOGIC METHODS. 

ing hemoglobin. This characteristic will readily serve to dif- 
ferentiate it from the colon bacillus and other organisms of sim- 
ilar appearance. An easily made and satisfactory medium is 
prepared by spreading a little fresh blood upon the surface of 
an ordinary agar slant and inoculating this with the infected 
material. This organism only develops on artificial media be- 
tween the temperatures of 26° and 43° C, growing best at body- 
temperature. Upon the blood-agar slant incubated at 37° C, 
there will develop minute, transparent, watery colonies that are 
without structure, somewhat resembling droplets of due. They 
are usually discrete, and show little or no tendency to coalesce. 

Staining. — One of the best methods of staining is with a 
dilute watery solution of Ziehl's carbol-fuchsin (the color of 
the solution should be pale red). This solution should be al- 
lowed to act for five minutes. This organism is decolorized by 
Gram/s method. 

A second method of staining is with Loeffler's methylene- 
blue (for preparation see Appendix). This stain should be 
allowed to act for five minutes. Then wash in water, mount, 
and examine for blue organisms. 

BACILLUS OF DIPHTHERIA. 

From the grayish white deposit on the fauces of a diph- 
theritic patient, prepare a series of cultures in the following 
way : — 

Have prepared a few tubes of Loeffler's blood-serum. Pass 
a stout platinum needle which has previously been sterilized into 
the membrane and rub it around there ; then being careful that 
it touches nothing else, rub it carefully over the surface of two 
tubes. The tubes are then immediately replugged and placed in 
the incubator. If the case be one of true diphtheria, the tubes 
will be ready for examination the following day. 

The blood-serum mixture is to be preferred to the ordinary 
plate method because the organism of diphtheria grows better 
on this medium than upon any other : it is also a differential 
method in a general sense, because other organisms do not grow 
well on Loeffier's serum, hence a luxuriant growth at the expira- 
tion of twenty-four hours should always be considered diph- 
theritic until proven otherwise. 



GONOCOCOUS OF NEISSER. 251 

Appearance. — After twenty-four hours the tubes presenl a 
characteristic appearance. Their surfaces are marked by more 
or less irregular patches of white' or cream-colored growths, 
which is usually more dense at the center than at the periphery. 

Staining. — From this culture smears are made upon clean 
cover-slips or slides, dried and fixed in the usual way, and then 
stained with LoefHer's alkaline methylene-blue. There will now 
be seen, in a typical case, upon microscopic examination, slightly 
curved bacilli of irregular size and outline. In some cases they 
will be more or less clubbed at one or both ends; sometimes 
they are spindle shaped or may present curved edges. They are 
rarely or never regular in outline. Many of these irregular rods 
are seen to be marked at circumscribed points in which their 
protoplasm is deeply stained. This irregularity in outline and 
appearance is the morphologic characteristic of the bacillus of 
diphtheria. 

THE GONOCOCCUS OF NEISSER. 

On microscopic examination the pus from an acute case of 
gonorrhea will show numerous small bodies, usually arranged 
in pairs. These cells will be found both within the protoplasm 
of some of the pus cells. The cells containing the gonococcus 
are usually crowded with the organisms, though the majority 
of pus cells do not contain them. This organism, on account of 
its frequent arrangement in pairs, is often called the diplococcus 
of gonorrhea. It is always found in gonorrheal pus and often 
persists into the convalescent stage after the external discharge 
has disappeared. It is easily detected in the pus of acute in- 
vasion, while in the subacute and chronic conditions its detec- 
tion is often a matter of considerable difficulty. 

Cultivation. — It does not grow upon the ordinary culture 
media, and can only be isolated in culture through the employ- 
ment of special methods. Blood or blood-serum is a necessary 
constituent of all media for the artificial cultivation of this 
organism. Some investigators have been successful in growing 
it upon other body fluids such as ascites fluid, pleural effusion, 
and the fluid from ovarian cysts. A useful medium may be 
prepared by mixing equal parts of human blood-serum with ordi- 
nary sterilized nutrient agar-agar. This is accomplished by 



252 BACTERIOLOGIC METHODS. 

liquefying the agar maintaining it at a temperature of 50° C. 
until after the mixture is made, after which it is allowed to 
solidify. 

Distinguishing Characteristics. — 1. It is seen practically 
always in the form of a diplococcus, having the characteristic 
biscuit form with the long diameters of the individual cells 
apposed. 

2. In gonorrheal pus some organisms are practically always 
found within the protoplasm of some of the cells. 

3. It stains readily with the ordinary staining reagents, but 
it is promptly decolorized by Gram's. 

4. It fails to develop on the ordinary artificial media (sepa- 
rating it from the diplococcus intracellularis meningitidis which 
grows freely). 

5. It has no pathogenic properties for the lower animals. 
Staixixg. — The ordinary stains are satisfactory, one of 

the simplest being a 1- or 2-per-cent. aqueous solution of methy- 
lene-blue which gives a very clear picture. 

Most important as a differential method is its failure to 
retain its color when treated by Gram's method. 



SPECIAL STAINING METHODS. 
Gram's Method. 

Method. — Objects are first treated with an aniline water 
solution of gentian-violet which is made after the formula of 
Koch-Ehrlich (see Appendix for preparation of stain). . After 
staining in this solution for fifteen to thirty minutes the excess 
of stain is drained off and the film treated for five minutes with 
Gram's iodine solution (see Appendix). They are next trans- 
ferred to alcohol and thoroughly rinsed. If careful examina- 
tion at this point reveals any violet color in the film, it must 
again be treated with the iodine solution until all violet color is 
removed. After a final washing in alcohol the specimen may 
be mounted and examined or a counter-stain of carmine may 
first be employed. 

Wright's Modification. — Stain for one minute in carbol- 
gentian-violet (see Appendix). Wash in water from thirty 



GRAMS METHOD. 253 

to sixty seconds. LugoPs solution is then allowed to act upon 
the specimen for one to three minutes. Wash and dry. Dif- 
ferentiate with aniline-xylol (2:1) to which 1.5 per cent, of 
acetone has been added, for one or two minutes. Wash with 
xylol, dry, and counter-stain with dilute carbol-fuchsin (1: 10) 
for about one minute, during which the specimen should be 
warmed slightly. The specimen is finally washed, dried, and 
mounted for examination. 

The process of decolorizing is only a relative one, some 
bacteria decolorizing more readily than others, so that much 
depends upon the intensity of the decolorizing reagent, and also 
upon the time during which it is allowed to act. The counter- 
stain method with dilute carbol-fuchsin is a differentiated proc- 
ess indicating those organisms that have been decolorized. All 
decolorized organisms by this method take on a red color. This 
counter-stain is of particular value where pictures of a number 
bacteria are made, as in sputum examination. 

The following organisms retain the violet stain by Gram's 
methods : — 

Streptococci. 

Staphylococci. 

Bacillus tuberculosis. 

Bacillus anthracis. 

Bacillus aerogenes capsullatus. 

Bacillus diphtherias. 

Diplococcus meningitidis intracellularis. 

Diplococcus pneumoniae (FrankePs). 

Bacillus tetanus. 

The following organisms are decolorized by Gram's 
method : — 

Bacillus pyocyaneous. 

Micrococcus gonorrhea (Xeisser) . 

Bacillus malignant oedema. 

Bacillus influenza. 

Bacillus typhosis. 

Bacillus cholera. 

Bacillus pneumonia (Friedlander's). 



254 BACTERIOLOGIC METHODS. 

Loeffler's Method of Staining Flagella. 

It is essential that the bacteria be evenly and not too nu- 
merously distributed over the cover-slip. 

Preparation of Cover-Slip. — The glasses must be perfectly 
clean. Lay six cover-slips on an even surface, and place in the 
center of each a small drop of distilled water. From the mate- 
rial to be examined, transfer a minute quantity to the drop of 
water on the first cover-slip; from this one transfer a minute 
quantity to the second, and so on to the sixth. This insures a 
varying dilution of the organisms in the different preparation. 
They are then all spread, dried, and fixed in the usual way. 
The cover-slip preparations are next warmed in LoefHer^s mor- 
dant (see Appendix). 

A few drops of this solution is placed upon the preparation 
which is held over a low Bunsen flame until it begins to steam. 
It should not be boiled. After steaming for a few moments the 
mordant is washed off with water and then with alcohol. The 
bacteria are now stained in the Ivoch-Ehrlich aniline water- 
fuchsin solution (see Appendix). 

When treated in this way various bacteria behave differ- 
ently, the flagella of some staining readily, others require the 
addition of an alkali in varying proportion to obtain the best 
results, others again stain best after the addition of an acid. 

To meet these conditions an exact 1-per-cent. solution of 
caustic soda in water must be prepared, and also a solution of 
sulphuric acid of such strength that 1 cubic centimeter will 
exactly neutralize 1 cubic centimeter of the alkaline solution. 

For the different bacteria which have been studied by this 
method Loeffler recommends that one or the other of these solu- 
tions be added to the mordant before using, in the following 
proportions : — ■ 

Of the Acid Solution. 

For Spirillum concentricum. no addition of either acid or alkali. 
For Spirillum cholera Asiaticae, % to 1 drop to 16 cubic centimeters 

of the mordant. 
For Spirillum Metchnicovi, 4 drops of acid to 16 cubic centimeters 

of the mordant. 
For Spirillum rubrum. 6 drops of acid to 16 cubic centimeters of 

the mordant. 



STERILIZATION. 255 

For Bacillus pyocyaneus, "> drops of acid to 16 cubic centimeters 
of the mordant. 

Of the Alkaline Solution. 

For Bacillus mesentericus vulgaris, 4 drops of alkali to 16 cubic 
centimeters of mordant. 

For Bacillus Micrococcus a<ji]is. 20 drops of alkali to 16 cubic centi- 
meters of mordant. 

For Bacillus typhosus, 22 drops of alkali to 16 cubic centimeters of 
mordant. 

For Bacillus subtilis, 28 to 30 drops of alkali to 16 cubic centi- 
meters of mordant. 

For Bacillus malignant edemse, 36 to 37 drops of alkali to 16 cubic 
centimeters of mordant. 

For Bacillus symptomatic anthrax, 35 drops of alkali to 16 cubic 
centimeters of mordant. 

The drops used run 22 to the cubic millimeter. 

STERILIZATION. 

General Considerations. — Acquaintance with the funda- 
mental principles of sterilization and of disinfection are abso- 
lutely necessary to the successful performance of all baeterio- 
logic investigations. The term sterilization, as commonly em- 
ployed, implies the absolute destruction of bacterial life by 
heat, while the term disinfection is commonly applied to accom- 
plishing the same end through the agency of chemical sub- 
stances capable of destroying bacterial life. 

Strictly speaking, the term sterilization implies the com- 
plete destruction of the vitality of all micro-organisms that may 
be present in or on the substance or substances to be sterilized. 
Such a result can obviously be accomplished by either thermal 
or chemical means, while disinfection need not, of necessity, de- 
stroy all living organisms that are present, but only those having 
the power of infecting or of producing disease, and may or may 
not, as the case may be, cause complete destruction of bacterial 
life, as in sterilization. It is therefore possible to accomplish 
both disinfection and sterilization by either chemical or thermal 
means. 

In the laboratory the employment of these different terms 
depends upon and is governed by circumstances. It is, of course, 
essential that all culture media should not onlv be absolutelv 



256 BACTERIOLOGIC METHODS. 

free from all bacteria, whether they are pathogenic or not, but 
also from their spores. In a word, they must be sterile. At the 
same time it is equally essential that the original chemical com- 
position and physical properties of the media should remain un- 
altered by the process. It is self-evident, therefore, that steriliza- 
tion of such substances by means of chemical agents, is out of 
the question, for while this method would destroy all bacterial 
life, it would not only alter the chemical composition of the 
media, but by becoming inseparably mingled with the media, 
would, by its continued presence, effectually prevent the growth 
of bacteria in the material for all time; that is to say, after 
having performed its function as sterilizer, it would by its con- 
tinued presence exercise its function as an antiseptic, and render 
the material useless as a culture medium. Exceptions to this 
general rule are found in certain volatile substances such as alco- 
hol and ether which, after having performed their bactericidal 
powers, may be completely driven off by the application of heat. 

Sterilization by Heat. — Sterilization by means of high tem- 
peratures may be accomplished in a variety of ways : 1. By dry 
sterilization, which is accompanied by subjecting the articles to 
adequate degrees of heat in a properly constructed oven. 2. 
By subjecting them to the influence of live steam at 100° C. 3. 
By subjecting the substances to steam under pressure. When 
employing steam under pressure, the temperature to which the 
articles are subjected will depend upon the pressure developed — 
the greater the pressure the higher the temperature. 

Sterilization by Dry Heat. — This method has the following 
disadvantages which limit its applicability : 1. The temperature 
must be relatively high and the period of exposure long as com- 
pared to moist heat (steam). 2. The penetration of dry heat 
into substances to be sterilized is much less thorough than that 
of steam. 3. Many substances of vegetable and animal origin 
are rendered valueless by the temperature required for dry ster- 
ilization. 

Successful sterilization by dry heat cannot usually be accom- 
plished by a temperature lower than 150° C, and this tempera- 
ture must be continued for not less than an hour. In general, it 
may be said that dry sterilization is only suited for sterilization 
of such substances as glassware, dishes, flasks, test-tubes, pi- 



STERILIZATION. 257 

pettes, etc., and for such metal instruments as are not injured 
by heat. 

Steam or moist heat sterilization possesses great penetrating 
power, and is much more rapid and thorough than the above 
method, and, further, it is far less likely to destroy the material 
so treated. This method should be employed for sterilizing all 
culture media, fabrics, cotton, wood, and organic material in 
general. 

Aside from the relative applicability of the two methods, 
their mode of action toward the organisms to be destroyed is 
very different. The penetrating power of steam renders it far 
more efficacious than dry heat. The spores of several organisms 
which are destroyed by exposure to steam for a few minutes, 
resist the destructive action of dry heat at a higher temperature 
for a longer period of time. 

The method of applying heat for sterilization depends 
chiefly upon the character of the substances to be sterilized. The 
application of dry heat is always continuous, i.e., the objects to 
be sterilized are simply exposed to the proper temperature for 
the requisite time necessary to destroy all living organisms and 
their spores which are either in or upon them. With steam, on 
the other hand, the articles to be sterilized are frequently of 
such a nature that prolonged sterilization would injure them. 
For this reason it has been found desirable to subject such ob- 
jects to the influence of steam intermittently for a number of 
short periods. 

The peixciple involved in the intermittent method de- 
pends on the differing powers of resistance to heat displayed 
by different organisms in different stages of their development. 
During the life of many bacilli they enter a stage during which 
their resistance to both chemical and thermal agents is mate- 
rially increased. This increased power of resistance is possessed 
by the organisms when they are in the spore or resting stage. 
Some spores of certain organisms have been encountered which 
retain the power of germination after an exposure of more than 
an hour to the temperature of boiling water. This difference in 
the thermal heat-point of bacteria and their spores is taken 
advantage in the process of sterilization known as the fractional 
or intermittent method. 

17 



258 BACTERIOLOGIC METHODS. 

As all culture media depend for their usefulness upon more 
or less unstable organic compounds, the effort of sterilization is 
to destroy the organisms in the shortest possible time by expo- 
sure to least possible amount of heat. This is accomplished by 
subjecting them to a temperature at a time when the bacteria 
are in the vegetative or growing stage. In order to develop any 
existing spores the media, during the intervals between steriliza- 
tion, should be kept under such conditions of temperature and 
moisture as will favor the process of vegetation (room-tempera- 
ture). 

During the first application of heat the mature vegetative 
forms are destroyed, while certain spores which may be present 
resist this treatment and survive the temperature. Xow the 
sterilization is discontinued and the media is allowed to remain 
for a time, usually twenty-four hours at room-temperature. 
During this time those spores which resisted the first heating 
have conditions favorable to germination. A second short ex- 
posure kills this crop of bacilli, when a second rest, followed by 
a third short exposure, kills the remaining organisms and the 
media will usually be found sterile. 

It should be remembered that while all spores which are 
present are not killed by the first exposure, still their power of 
germination may be so inhibited by the exposure to 100° C. 
that their germination is delayed, that they cannot possibly 
germinate during the twenty-four hours' intervals. 

Experiment has shown that the fractional process gives the 
best results when the objects are subjected to the action of live 
steam (steam at ordinary atmospheric pressure) for fifteen 
minutes on each of three consecutive days, and that during the 
intervals the cultures should be maintained at a temperature 
between 25° and 30° C. The substances thus treated will re- 
main sterile for an indefinite time, provided they are not exposed 
to the re-entrance of micro-organisms. 

An occasional exception will be noted when, after careful 
treatment as above outlined, certain species of spore-forming 
bacteria will not have been entirely destroyed by this method. 
These are usually of the non-pathogenic group of the so-called 
soil organisms. 

Finally it must be born in mind that this method is only 



STERILIZATION. 259 

applicable to substances capable of presenting conditions favor- 
able to spore germination, and that dry substances, such as in- 
struments, apparatus, or organic materials, in which decomposi- 
tion has set in, where the natural conditions favorable to spore 
germination are absent should not be treated by this method, 
but must be subjected to higher temperatures for longer periods 
of time. 

Intermittent Sterilization (at low temperature). — The 
process of intermittent sterilization at comparatively low tem- 
peratures is based upon the principle outlined above, but differs 
in the details of its application as follows : 1. It requires a 
greater number of exposures to accomplish complete sterilization. 
2. The temperature at which it is accomplished is not above 
68°-70° C. 

It is employed for sterilization of easily decomposable ma- 
terials and those which would be rendered useless by the appli- 
cation of steam at 100° C, but which are unaltered by the tem- 
perature employed. This method is applicable for sterilization 
of certain albuminous media where it is desirable to retain 
their fluid condition during sterilization and which would be 
coagulated by exposure to higher temperature. 

This process requires that the temperature employed should 
be between 68° -70° C, and that an exposure of one hour should 
be made each of six consecutive days. During the intervals the 
material is kept at a temperature between 25°-30° C. to favor 
germination of any spores that may be present. 

Direct Steam Method. — Sterilization by means of steam is 
also accomplished by what is known as the direct or continuous 
method. By this process both mature organisms and their spores 
are destroyed by a single exposure to steam at zero pressure — 
live or steaming steam. The sterilization is accomplished by a 
single exposure of one hour. 

Steam-Pressure Method (by Autoclave). — By employing 
steam under pressure we are able, by increasing the temperature, 
to materially shorten the time necessary to accomplish complete 
sterilization. By employing a pressure of approximately one 
atmosphere (15 pounds) a temperature of about 122° C. is 
obtained. This is sufficient to accomplish complete sterilization 
by one exposure of fifteen minutes. 



260 



BACTERIOLOGIC METHODS. 



When this method is employed it will occasionally be found 
that the coagulating power of gelatin is reduced, and that it 
becomes slightly cloudy, while in agar-agar a fine, flaky precipi- 
tate is noticed. For accurate time and temperature exposures 
this is a very uncertain method. Obviously the material is sub- 
ject to active temperatures during the heating up as well as 
during the cooling process, besides the actual time during which 
the maximum pressure is maintained. Also, if great care is not 




Fig. 31.— Arnold Steam Sterilizer. (A. H. Thomas.) 



observed to prevent premature opening of the autoclave, a 
sudden, rapid ebullition of the fluid occurs which, in the case 
of test-tubes, the media may completely boil away. 

While this method of sterilization is not well suited for 
delicate experiments where a definite time exposure to definite 
temperature is of importance, still, for general laboratory pur- 
poses, it has much to recommend it in the way of a time saver, 
and in the certainty with which sterilization is accomplished. 
The apparatus designed to sterilize under pressure is termed an 
autoclave. 



THE AUTOCLAYK. 



261 



Practical Application of the Method. — 1. Tin: Arnold 
Steam Sterilizer (Fig. 31): This is probably the best steril- 
izer for general laboratory purposes, since it is simple and eco- 
nomic in its operation. The difference between this apparatus 
and the original sterilizer devised by Koch is that it provides 
for the condensation of the steam after its escape from the 




Fig. 32.— Autoclave. (A. H. Thomas.) 



sterilizing chamber, and returns the water of condensation auto- 
matically to the reservoir. 

2. The Autoclave. — The advantage of this method is so 
well recognized that its use has practically superseded the inter- 
mittent method w T ith live steam, except when the temperature 
developed by the autoclave is sufficient to destroy the materials 
subjected to it. By this plan sterilization is accomplished in 
fifteen minutes by exposure to steam under the pressure of one 
atmosphere. 



262 BACTERIOLOGIC METHODS. 

The autoclave (see Fig. 32) embodies the same principle as 
the steam sterilizer. It provides for the generation of steam 
within a chamber capable of being hermetically sealed after the 
introduction of the substances to be sterilized. The chamber is 
fitted with a safety-valve arranged for regulating the degree of 
pressure. A thermometer passes through the wall and enters 
the chamber, thus allowing the temperature to be followed and 
the pressure properly regulated. 

Hot-Air Sterilizer. — The hot-air sterilizers employed for 
this work are simply double-walled boxes of Swedish iron, having 
a double door and a copper bottom. They are fitted with proper 
openings in the walls to permit circulation of the heated air. 
The heat is obtained from the flame of a Bunsen burner applied 
directly to its bottom. These bottoms are usually constructed of 
copper, and because they readily burn out, are now made so that 
they may easily be replaced. Properly constructed sterilizers 
with removable bottoms may now be obtained and should be 
sought when purchasing, as they will save much annoyance and 
not a little expense. 

Sterilization by this method is accomplished in from a half 
to one hour at a temperature of from 150°-180° C. 

CHEMICAL STERILIZATION AND DISINFECTION. 

It is possible by means of certain chemical substances to 
destroy all bacteria and their spores, or by the same means to 
remove all their pathogenic properties; in other words, to dis- 
infect them. 

When it is desirable to use chemical disinfection in the 
laboratory, successful results may be obtained by employing a 
33- to 40-per-cent. solution of carbolic acid. Under ordinary 
circumstances this will accomplish the result in from twenty 
minutes to half an hour. It is, however, not reliable for the 
destruction of spores of resistant organisms, such as the spores 
of the anthrax bacillus. 

All materials and issues containing infectious organisms 
should be burned and all other cloths, test-tubes, flasks, dishes, 
etc., should be boiled in a 2-per-cent. soda (ordinary washing 
soda) for a half-hour, or should be exposed in the steam ster- 
ilizer for the same length of time. 



PREPARATION OF CULTURE MEDIA. 263 

Intestinal evacuations are best disinfected with chlorinated 
lime, which should contain at least 0.25 per cent, of free chlo- 
rin. This solution should be mixed in equal parts with the 
material to be disinfected, and then should be allowed to stand 
for one or two hours before being disposed of. 

Sputum in which tubercle bacilli may be present, as well 
as vessels containing it, and the eating utensils of tuberculous 
patients, should be boiled with a 2-per-cent. soda solution for 
from a half to one hour, or should be exposed in the steam ster- 
ilizer for the same period. 



PREPARATION OF CULTURE MEDIA. 

Bouillon. — Five hundred grams of freshly chopped, lean 
beef, free from fat and tendons, are soaked in a liter of water 
for twenty-four hours, during which time the temperature of the 
mixture is kept low by surrounding it with ice. 

At the expiration of this time the mixture should be strained 
through coarse muslin until a liter of fluid has been recovered. 
To this now add ten grams of dried peptone and five grams of 
common table salt. It is then to be rendered neutral or slightly 
alkaline by the addition of a few drops of a saturated sodium 
carbonate solution. The flask containing the mixture is then 
placed either in the steam sterilizer or upon a water-bath and 
kept at the boiling point until all the albumin has been coagu- 
lated, and the fluid portion is clear and of a pale-straw color. 
It is then filtered through a folded filter-paper and finally ster- 
ilized in the steam sterilizer by the fractional method (see page 
259). This is the original method of Koch which has been 
modified and improved in the following ways: — 

Neutralization. — Ordinarily this is accomplished by the 
addition of a saturated solution of sodium carbonate, and the 
reaction determined by red and blue litmus paper. This 
sodium carbonate solution is not so good, however, as a strong 
solution of sodium or potassium hydroxid, because the carbonic 
acid arising during the process of neutralization with the sodium 
carbonate frequently produces a temporary acid reaction which 
later disappears on boiling. Exact titration with an 0.4-per-cent. 
solution of sodium hydrate, obviates this difficulty. The process 



264 BACTERIOLOGIC METHODS. 

is applied only after the bouillon has been deprived of all its 
coagulable albumin by boiling and has again been reduced to 
room-temperature. 

Techxic. — First ascertain the exact volume of the fluid. 
From this sample take exactly 5 or 10 cubic centimeters and 
add a few drops of a 1-per-cent. alcoholic solution of phenol- 
phthalein as an indicator. The O.-4-per-cent. alkaline solution 
is placed in a graduated burette, and the solution to be tested 
in a porcelain dish or casserole. Now add the alkaline solution, 
drop by drop, until the bouillon turns a faint rose color. A 
second measured quantity of bouillon is treated in the same 
manner as a check, and if the amount of sodium hydrate solu- 
tion required to cause neutralization is the same or only slightly 
different, a simple calculation will indicate the amount of soda 
solution required to neutralize the bulk of the medium. Thus, 
if for 10 cubic centimeters of the bouillon would be required 1.5 
cubic centimeters of the O.-1-per-cent. solution of sodium 
hydrate, then for the remaining 980 cubic centimeters (original 
volume 500 cubic centimeters), it would require 98 times 1.5 
cubic centimeters of the soda solution to neutralize the total 
amount of bouillon, or 14:7 of the O.-1-per-cent. sodium hydrate 
solution would have to be added to the 980 cubic centimeters of 
bouillon to accomplish neutralization. 

To avoid over-diluting the bouillon by the weak alkaline 
solution, it is better to employ for this purpose a 4.0-per-cent. 
solution of XaOH, of which only 1-4.7 cubic centimeters would 
be required. 

Xot infrequently the filtered neutralized and sterilized 
bouillon will be found to contain a fine flocculent precipitate. 
This may be due either to an excess of alkalinity or to incom- 
plete precipitation of the albumin. The former may be corrected 
by the addition of a little dilute acetic or hydrochloric acid, fol- 
lower by a second boiling, filtering, and sterilization. If due to 
imperfect precipitation of the albumin this may be corrected by 
reboiling and filtering. 

2. The substitution of prepared meat extract for the fresh 
extract is now almost universal. Any good stock meat extract 
will answer the purpose, and should be used in the strength of 
from two to four grams to the liter of water. Peptone and 



PREPARATION OF CULTURE MEDIA. 265 

sodium chlorid are added, as in the original method of prepara- 
tion. 

The advantages of the meat extract are a decided shorten- 
ing of the time required for preparation of the media, and the 
production of a more uniform material. 

Nutrient Gelatin. — In making nutrient gelatin the bouil- 
lon is prepared first as outlined above, except that the reaction 
is corrected only after the gelatin has been completely dissolved. 
The reaction of the gelatin of the shops is frequently quite acid, 
entailing the addition of considerable more alkaline solution 
than required for the bouillon alone. 

The gelatin is added in sufficient quantity to make a 10- 
or 12-per-cent. solution. Its complete solution is accomplished 
either on a water-bath or over the bare flame. In the latter 
instance it must be constantly stirred to prevent burning the 
gelatin on the bottom of the pan. When the gelatin is com- 
pletely dissolved it can readily be filtered through an ordinary 
folded filter paper in a glass funnel. It not infrequently happens 
that in spite of the most careful technic the filtered gelatin is 
not perfectly clear; then clarification is required. For this pur- 
pose the mass must be redissolved, and when the temperature is 
between 60° and 70° C. the white of an egg, which has been 
beaten up with about 50 cubic centimeters of water, is added 
and thoroughly mixed in. This mixture is then again brought 
to the boiling point until coagulation of the egg albumin occurs. 
This process results in large, flaky coagula of albumin, which it 
is best not to break up, as fine flakes of albumin will clog the 
filter-paper and materially interfere with the process of filtra- 
tion. 

The filter-paper should always be moistened before filtering. 
If this is not done, the pores of the filter-paper will become 
clogged with gelatin and coagulated albumin, which will greatly 
interfere with rapid filtering. 

Gelatin should not, as a rule, be boiled for more than fifteen 
minutes, nor left in the steam sterilizer for more than thirty 
minutes, otherwise its property of solidifying will be impaired. 

As soon as the preparation of the gelatin is completed it 
should be sterilized in the steam sterilizer for fifteen minutes on 



266 BACTERIOLOGIC METHODS. 

three consecutive days. The mouths of the containing flask or 
test-tubes should be completely plugged with raw cotton. 

Nutrient Agar-Agar. — The preparation of this is difficult 
and tedious, and frequently fails from lack of patience or of 
experience, or both. Every worker has some slight modification 
of his own, but if, according to Abbott, the following directions 
are carefully carried out, the product will usually be satis- 
factory. 

Prepare bouillon in the usual way. As agar-agar re- 
acts neutral or faintly alkaline, the neutralization of the bouil- 
lon may be accomplished before the agar-agar is added. Finely 
chopped or powdered agar-agar is added to the bouillon in the 
proportion of 1 to 1.5 per cent. The mixture is then placed in 
an agate or earthen-ware boiler, and the height of the fluid, 
before boiling, marked upon its inside. If a liter of the 
medium is being made add about 275 cubic centimeters or more 
of water, and boil slowly for about two hours or until the excess 
of water that was added has been evaporated. Do not allow the 
fluid in the vessel to fall below the original level. If this occurs 
water must be added to bring the final amount up to one liter. 
At the expiration of two hours remove from the fire and cool 
rapidly by immersion in a pan of cold water. Stir constantly 
until the temperature of the mass has fallen to about 70° C, 
then add the white of one egg that has been previously beaten 
up in about 50 cubic centimeters of water. Mix this in well and 
allow to boil for half an hour longer, keeping the fluid up to the 
original one liter mark. The fluid is now easily filtered at 
room-temperature through a heavy folded filter. If properly 
prepared and filtered through a properly folded filter-paper, it 
should pass through at the rate of one liter in from fifteen to 
twenty minutes. 

Glycerin Agar-Agar. — The nutrient properties of agar- 
agar for certain organisms is greatly increased by the addition 
of 5-per-cent. glycerin. If glycerin is added to the agar-agar, 
it should be done after filtration and before sterilization. 

If after filtration the medium is found to contain flocculi, 
investigate the reaction. If it is quite alkaline it must be neu- 
tralized, boiled, and filtered again. If the reaction is neutral 



PREPARATION OF CULTURE MEDIA. 267 

or only faintly acid, dissolve and again clarify with egg albumin 
as directed above. 

The most important feature of all media apart from then- 
proper preparation is the reaction. They must be neutral or 
only very faintly alkaline to litmus, as but few organisms de- 
velop well on acid media. 

Blood-Serum. — For the preparation of a small amount of 
blood-serum for culture media purposes, it may be obtained 
from small animals under such aseptic precautions as will guard 
against gross contamination. For laboratory purposes, where 
large amounts are required, it is best obtained from the slaughter 
houses. Under these conditions a certain amount of contamina- 
tion is unavoidable, though its extent may be limited by observ- 
ing certain precautions. 

The original method of Koch with a few slight variations 
is as follows: — 

The animal from which the blood is to be obtained should 
be suspended by the hind legs so that its head is a few feet from 
the floor. The head should be held back when, with one sweep 
of a sharp knife, the throat is completely cut through. The 
blood as it spurts from the vessels should be collected in large 
glass jars which have been previously sterilized and dried with 
alcohol and ether. The jars should be provided with close- 
fitting cover and clamps capable of hermetically sealing them 
(large museum specimen jars will be found very satisfactory for 
this purpose). From two-gallon jars of blood there is usually 
recovered from 500 to 700 cubic centimeters of clear serum. 

The jars having been filled with blood, their covers are re- 
placed loosely and then they are allowed to stand quietly for 
about twenty minutes until clotting has begun. At the expira- 
tion of this time a clean glass rod is passed about the edges 
of the surface of the forming clot to break up any adhesions 
that have formed to the sides of the jar. The covers are now 
replaced and clamped clown tightly, then with as little agitation 
as possible the jars are transferred to an ice chest where they 
should remain for from twenty-four to forty-eight hours. The 
temperature of the ice chest should be only sufficiently low to 
prevent bacterial growth, but not so cold as to prevent coagula- 
tion. When the jars are removed from the ice chest a firm clot 



268 BACTERIOLOGIC METHODS. 

will be found in the bottom of the jars. The serum is drawn 
off with a sterile pipette or syphon, and transferred to sterile 
glass cylinders. These cylinders are now placed on ice for an- 
other twenty-four hours, when the corpuscles will have settled 
to the bottom, leaving the serum above quite clear. This is then 
ready to be pipetted off into sterile test-tubes, about 8 cubic cen- 
timeters in each tube, or into flasks of 100 cubic centimeters' 
capacity. It is now ready for sterilization. This is accomplished 
by the intermittent method at a temperature of 70° C. for a 
period of one hour on five consecutive clays. During the intervals 
it should be kept at room-temperature. After sterilization the 
tubes may be allowed to remain fluid or they may be coagulated 
by a short exposure to 80° C. For solidifying, the tubes should 
be placed in an inclined position in order to secure the greatest 
possible surface from the quantity of serum employed. 

The process of solidification requires constant attention if 
good results are to be obtained. Xo rule can be laid down for 
the time required to accomplish it, as this is not constant. Too 
high temperature and too rapid solidification results in an opaque 
and inelastic medium. 

When solidification is complete the tubes may be retained 
in a vertical position, and unless for immediate use must be 
protected from drying. This may be done by burning off the 
superfluous ends of cotton and covering them with sterile rubber 
caps; or what is just as satisfactory and far cheaper, sterilized 
corks may be pushed down upon the cotton plugs. 

Owing to the employment of large quantities of serum, 
principally for the detection of diphtheria, the tedious method 
of Koch has been largely superseded by a number of more 
rapid modifications. 

Method of Councilman and Mallory. — By this method the 
serum is more quickly prepared. Eigid precautions against con- 
tamination of the blood during collection are not necessary, and 
the resulting medium, while neither transparent or translucent, 
fully meets the ordinary requirements of bacteriology. 

By this method the serum is decanted directly into sterile 
test-tubes as soon as obtained; it is then firmly coagulated in 
the slant position by exposure in a dry-air sterilizer at from 80° 
to 90° C. It is then immediately sterilized in the steam sterilizer 



PREPARATION OF CULTURE MEDIA. 269 

at 100° C. for fifteen minutes on three consecutive clays, as is 
the case with other media. 

Loeffler's Blood-Serum Mixture. — This mixture consists of 
one part neutral meat-infusion bouillon, containing 1 per cent, 
of grape-sugar and three parts blood-serum. The mixture is 
placed in test-tubes, sterilized, and solidified in exactly the same 
manner as described under blood-serum preparation, except that 
it requires a longer time and a higher temperature for coagula- 
tion. 

PREPARATION OF TUBES, FLASKS, ETC., FOR 
CULTURE MEDIA. 

While the media are in the course of preparation it is well 
to get the tubes, flasks, pipettes, etc., ready for their reception. 
These must be absolutely sterile. To this end both old and new 
tubes should first be boiled for a half-hour in a 2-per-cent. soda 
solution, then carefully swabbed out with appropriate bristle 
brushes. After rinsing in clear water they are immersed in a 
1-per-cent. solution of hydrochloric acid for a few minutes, then 
rinsed again and stood, round end up, to drain. When dry they 
are plugged with raw cotton carefully inserted so that there are 
no cracks or openings in the occluding cap. The plug should fit- 
neither too loosely nor too tightly, but should fit firm enough to 
hold the weight of the tube when lifted by the protruding cotton. 

The tubes thus plugged are then placed upright in wire 
baskets and heated for one hour in the hot-air sterilizer to a 
temperature of 150° C. Tubes so prepared, if undisturbed, will 
remain sterile for an indefinite period. 

Filling Tubes. — The tubes are best filled with the aid of 
a separating funnel, though if not convenient the tubes can, 
with a little care, be successfully filled directly from the flasks. 
It is not necessary to sterilize the funnel as the media in the 
tubes is to be sterilized as soon as they are filled. In any case, 
care should be observed to prevent any of the medium from 
coming in contact with the mouth of the tubes, which would 
cause the cotton plugs to adhere to it, making them hard to 
remove, presenting a very untidy appearance and materially in- 
terfering with subsequent manipulations. 

After filling, the tubes are ready for final sterilization. This 



270 BACTERIOLOGIC METHODS. 

is accomplished in the steam sterilizer by the three-day frac- 
tional method. 

TECHNIC FOR PLATES AND PETRI DISHES. 

Plates. — The plate method can be employed with both agar 
and gelatin, but cannot be practiced with blood-serum, because 
the latter when once it is solidified cannot again be rendered 
liquid. 

Plates are usually referred to as a set. This term includes 
three separate plates each representing a mixture of the organ- 
ism in a state of greater dilution. The plates are numbered 
1, 2, and 3. A set of plates may be prepared as follows : Three 
tubes, each containing the requisite amount of gelatin or agar- 
agar, are placed in a water-bath and warmed until the medium 
is fluid. Agar-agar becomes fluid at about the temperature of 
boiling water; gelatin is fluid between 35° and 40° C. In the 
case of the agar-agar tubes, after liquefying they must be 
cooled to 40° C, at which temperature they remain fluid while 
the organisms are introduced. If this cooling is omitted the tem- 
perature of the medium when the organisms are inoculated will 
be sufficiently high to destroy their vitality. 

The medium now being liquid and of a proper temperature, 
the material containing the organisms is taken up on a sterile 
platinum wire-loop and transferred to tube 1, where it is thor- 
oughly disintegrated and mixed by rubbing against the side of 
the tube. The more carefully this is clone the more uniform will 
be the distribution of the organisms and the better the final re- 
sults. The loop is now again sterilized by passing through the 
flame, and when cool three loops full from tube 1 are trans- 
ferred to tube 2, where they are carefully stirred in. Again, 
the wire is sterilized and the same manipulation carried out be- 
tween tubes 2 and 3. This completes the dilution. 

During these manipulations, which must be done rapidly if 
agar is employed, the temperature of the water bath must be 
kept between 39° and 43° C. If the temperature falls below 
38° C. the agar-agar will become solidified and can then only 
be reliquefied by the application of heat sufficient to destroy the 
organisms introduced. 

After inoculation the contents of these tubes are poured out 



THE INCUBATOR. 271 

upon the sterilized plates, cooled, and incubated for twenty-four 
or forty-eight hours. 

Petri Dish Method. — This process materially simplifies the 
original technic of the plate method. It consists in substituting 
for the flat plates of glass small, round double-glass dishes having 
about the same surface area as the original plates. The inocu- 
lated and liquid media is poured directly into these, their covers 
are immediately replaced, and they are set aside to cool. In all 
other respects the process is the same as Koch's original plate 
method. These dishes have vertical sides which prevents over- 
flowing of the medium. A convenient size for this method meas- 
ures about 12 centimeters in diameter, but dishes of other 
sizes are easily obtainable. The dishes are readily sterilizable 
by hot air or steam, and have the great advantages that the 
danger of contamination is reduced to a minimum, since after 
sterilizing the plates do not have to be separated until the pour- 
ing on of the medium, and then only for a moment. 

THE INCUBATOR. 

After the plates have been made and solidified they should 
be transferred to an incubator where a uniform and favorable 
temperature may be maintained. Various types of incubators 
have been devised, but since the principle and purpose of all is 
the same, a general description of their construction and method 
of employment is all that is required. 

The incubator or thermostat (Fig. 33) consists essentially 
of a copper chamber of convenient size provided with double 
walls, between which heated water circulates. The incubator 
chamber has a close-fitting door of heat-proof construction, and 
usually within this is a second door provided with a glass front, 
which permits inspection of the interior of the incubator with- 
out actually opening the chamber, and so reducing the tem- 
perature. The whole apparatus is set upon an enclosed base 
and is covered with asbestos board to prevent loss of heat from 
radiation. In the top of the chamber is a small opening fitted 
with a perforated cork through which a thermometer projects 
into the interior. Two other openings are provided, one for the 
thermometer which records the temperature of the circulating 
water, the other for the thermo-regulator. At one side of the 



272 



BACTERIOLOGIC METHODS. 



apparatus is a vertical water gauge provided with an upper 
opening for the introduction of water, and a stop-cock below 
for drawing off the water when occasion required. 

When in operation the apparatus should be kept full of 
water, otherwise the object of the water jacket will be defeated 
and the temperature of the interior of the chamber will not be 
maintained. Heat is supplied to the incubator by a gas burner 




Fig. 33.— Thermostat or Incubator. (A. H. Thomas.) 



placed within the inclosed space below the chamber. The par- 
ticular form of burner usually employed is known as "Koch's 
safety burner/' which is so constructed that should by accident 
the light be extinguished, the flow of gas would be almost im- 
mediately shut off. An ordinary Bunsen burner, well protected 
from sudden gusts of air, will serve the purpose equally as well. 
The Thermo-Regulator. — The efficiency of the thermostat 
depends upon the proper and uniform temperature which is 



THE INCUBATOR. 273 

maintained by the thermo-regulator. A satisfactory regulator 
should permit of a fluctuation of not more than 0.2° C. in the 
temperature within the chamber of the apparatus. 

The commonest form of regulator is constructed upon the 
principle involving the expansion and contraction of fluids under 
the influence of heat and cold. By means of such expansion and 
contraction the amount of gas passing from the source of sup- 
ply to the burner is modified, as the temperature of the water in 
the incubator rises or falls. 

For the successful employment of clinical laboratory meth- 
ods, the thermo-regulator is useful but not essential. In a room 
of fairly uniform temperature the flow of gas can be regulated 
by hand until the internal temperature of the incubator is 37° 
C. After this, if the water level is maintained and the apparatus 
protected from protracted changes in the surrounding tempera- 
ture, the variation in the internal temperature will be so slight 
that it need not be considered. 



18 



XVI. 

APPENDIX. 

DESCRIPTION OF OFFICE LABORATORY CABINET. 

For those who are daily employing the methods of clinical 
medicine a detailed description of the author's laboratory cabinet 
may be of interest. As illustrated in frontispiece, this cabinet is 
constructed of white pine or poplar, of half -inch boards, except 
the lowest shelf or table which is one-inch in thickness. This 
can easily be constructed by a carpenter and finished in white 
enamel. 

Its adoption will be found particularly useful to those not 
fortunate enough to have a room to devote to laboratory 
purposes. The cabinet itself is not at all unsightly, and may 
be fastened to the wall in the consulting room, preferably near 
a washstand, as running water should be easily accessible when 
possible. For those who possess the advantage of a well-equipped 
laboratory, it will be found extremely convenient for preliminary 
rough examinations of specimens as soon as they are received. 

The cabinet occupies a wall space of 22 ins. broad by 28 
ins. high, and projects outward from the wall 18 ins. The 
shelves are 6% ins. deep, the top shelf providing space for a 
gallon bottle of distilled water with syphon-tube attachment, 
and place for storage of large stock reagent bottles, beakers, 
flasks, dishes, etc. The compartment furnished with a glass door 
is for the microscope and accessories, where they may be safely 
kept under lock and key. 

The shelf below the top provides space for 24 2-ounce re- 
agent bottles or a less number of larger ones. The drawer im- 
mediately below this holds filter paper, Petri dishes, forceps, 
short pipettes, etc., while the small compartment to its right is 
(274) 



OFFICE LABORATORY CABINET. 



275 



very convenient for a few test-tubes, cover-glasses, slides, and 
litmus paper. 

The table, which measures 18 by 21 ins., is sufficiently large 
to contain the necessary racks, stands, etc. Passing through the 
table to its extreme left is a gas-cock with nipple for the attach- 
ment of a Bunsen burner. 

Such a cabinet will provide all the necessities for the usual 
examinations of urine, gastric contents, and sputum. By modi- 
fication and amplification of this idea the cabinet can be adapted 
to all the special examinations employed in clinical medicine. 

EXACT DIMENSIONS OF CABINET. 

Total height, 28 ins.; total width, 22 ins.; depth of table, 
18 ins. ; depth of shelves, 6% ins. ; height of drawer base above 




Fig. 34.— Portable Urinalysis Set. 



table, 8 ins. ; size of drawer, 6 by 10 by 4 ins. ; microscope com- 
partment, 8 ins. wide by 14 ins. high. ; height from table to top 
shelf, 24 ins. 

The w r ood should be carefully selected, free from knots and 
thoroughly seasoned. It should be joined by screws and glue to 
prevent warping and separation of the joints and seams. Two 
coats of good white paint, followed by the same number of white 
bath-tub enamel, will complete the construction. The door and 
drawer-knobs are of glass, and the microscope compartment is 
fitted with a lock. 

Since the construction of this cabinet three years ago the 
following improvements have suggested themselves: A large 
shallow drawer could be placed under the table. This would 



276 APPENDIX. 

be very useful for storing burettes, large pipettes, etc., and would 
not materially add to the size of the piece. A wooden burette 
stand and filter holder could be easily attached to the under 
surface of the first shelf, and so arranged that when not in use 
they could be swung under the shelf and out of the way. 

Fig. 34 shows a portable urinalysis outfit, which is con- 
venient for bedside work. It may be obtained in the open 
market at a moderate price. 



FORMS OF CLINICAL REPORTS. 



277 



URINALYSIS. 

Report of Clinical Laboratory. 



Physical Characteristics. 



Quantity in | 
24 hours. J 
Color, 

Appearance, 
Odor, 
Sediment : — 

Mucus, 

Urates, 

Phosphates, 

L^ric Acid, 

Pus, 
Specific Gravity, 
Reaction , 



c.cm. 
oz. 



Quantitative Determinations. 

•'Heat, 10% Acetic Acid. 
Albumin J Heller's .Nitric Acid. 
LHeat and Nitric Acid. 

TFehling's. 
Glucose J Phenylhydrazin. 

LBottger's Bismuth. 

Indican, 

Skatol, 

Acetone, 

Bile Pigments, 

Blood, 

Diazo, 

Aceto-acetic Acid, 

B-Oxybutyric Acid, 

Drug Reactions, 

Cammidge Reaction, 



Quantitative Determination. 



Albumin ( Esbach ) , ( Purdy ) , 

Glucose (Fehling's), (Fermentation), 

Urea, 

Ethereal Sulphates, 

Chlorids, 



Microscopic Examination. 





(Centrifugated), 


(Sedimented). 




r Hyaline, 
Granular, 




Erythrocytes, 
Leukocytes, 




Fatty, 




Pus cells, 


Casts.-< 


Epithelial, 

Blood, 

Waxy, 




Epithelia, 

Spermatazoa, 

Bacteria, 




Bacterial, 




Yeast Spores, 


Cylindroids, 
Mucus, 




Trichomonides. 


Voided, A.M 


.P.M 


Examined. A.M P.M 


Exami 


ned bv 







2.78 



APPENDIX. 



SPUTUM EXAMINATION. 

Report of Clinical Laboratory. 









Physical Characteristics. 


Quantity in > 
24 hours. ) 


c.cm. 
oz. 


Color, 




Odor, 




Consistence, 




Blood, 




Character, 




Serous, 




Mucous, 




Muco-purulent, 
Purulent, 




Casts, 




Spirals, 






Microscopic Examination. 


Tubercle Bacilli, 


Blood-Cells, 


Micrococcus Lanceolatus, Pus Cells, 


Staphylococci, 


"Heart Failure" Cells, 


Streptococci, 
Bacilli, 


Hematin Crystals, 
Elastic Fiber, 


Actinomyces, 
Yeast, 


Curschmann's Spirals, 
Casts, 


Obtained 


Examined 


Examined by 





FORMS OF CLINICAL REPORTS. 279 

BLOOD EXAMINATION. 

Report of Clinical Laboratory. 



General Examination. 

Color, 

{Estimated, 
Wright's, 
Hogg's, 

Viscosity, 

Flow from Puncture, 

Specific Gravity, 

C Thoma-Zeiss per c.mm., 
Erythrocytes J Estimated by Daland Hematokrit, 
[_ Percent., 

C Fleischl's Hemoglobinometer, 
Hemoglobin J Sahli's Hemoglobinometer, 
[_ Tallqvist's Scale, 

Color Index, 

Plaques, 

Leukocytes, 

Microscopic Examination. 

Erythrocytes. 

Color, Macrocytes, 

Polychromatophilia, Microcytes, 

Granular Degeneration, Normoblasts, 
Poikilocvtosis, 



Leukocytes, 



Differential Count, 

Number of cells counted, 

Polymorphonuclears, 

Large lymphocytes, 

Small lymphocytes, 

Eosinophiles, 

Basophiles, 

Myelocytes, 

Parasites, 

Bacteria, 

Hematin Crystals, 

Date 

Examined by 



280 



APPENDIX. 



GASTRO-ANALYSIS. 

Report of Clinical Laboratory. 



Vomitus, 






Test meal 






Composition : — 






Ingested, 






Extracted, 






Amount Recovered, 




Dilution, 








Physical Chakacteristics. 






Sediment containing : — 


Color, 




Blood, 


Odor, 




Bile, 


Consistency, 




Mucus, 


Reaction, 








Chemical Determinations. 


Total Acidity, 




Butyric Acid, 


Free Hydrochloric Acid, 


Acetic Acid, 


Combined " 


a 


Blood, 


Lactic Acid, 




Bile, 


Protealysis, 








Microscopic Examination. 


Starch Grains, 




Oppler-Boas Bacillus, 


Meat Fiber, 




Sarcinse, 


Epithelia, 




Bacteria, 


Erythrocytes, 




Necrotic Tissue, 


Leukocytes, 




Parasites, 


Date 




. . . Examined 


Examined by 







FORMS OF CLINICAL REPORTS. 



281 



BLOOD-PRESSURE DETERMINATIONS. 

Clinical Report. 



ApparatusJ 

L Stanton, 




Width of Cuff 


cm. 


Part examined, 




Right, 




Left, 




Posture, 




Pulse Rate, 




Systolic 


mm.Hg. 


Diastolic 


mm.Hg. 


Pulse Pressure 


mm.Hg. 


Mean Pressure 


mm.Hg. 


Remarks. 





Time of Day. A. M P. M., 



Date 



Examined by 



282 



APPENDIX. 



EXAMINATION OF THE FECES. 

Report of Clinical Laboratory. 



Physical Characteristics. 
Number of Stools Mucus, 

in 24 hours. Parasites, 

Amount in 24 Hours. Crystals, 



Color, 

Odor, 

Consistence, 

Blood, 

Pus, 



Calculi, 

Connective tissue, 
Muscle fiber, 
Foreign bodies, 



Chemical Examination. 



Blood, 

Hydrobilirubin (sublimate test), 

Fermentation, 

"Lost" Albumin. 



Erythrocytes, 
Pus cells, 
Epithelia, 
Crystals, 
Meat Fiber, 
Connective Tissue, 
Starch Cells, 
Free Fat, 
Fatty acids, 



Microscopic Examination. 

Tubercle Bacillus, 

Shiga Bacillus, 

Comma Bacillus, 

B. Aerogenes Capsulatus, 

Amebse, 

Parasites, 



Obtained 



Examined 



Examined by 



CLINICAL TERMS. 283 



CLINICAL TERMS. 

Owing to the various and uncertain terms employed to 
designate the amounts of substances found, in clinical investiga- 
tions the following scheme has been successfully employed in 
our laboratory. 

Terms to be used in expressing the results of examinations 
of specimens in laboratory 1 : — 





r Questionable trace. 


A 1 VvTl TVI "1 T"l 


Very faint trace. 


j^lOUIIllIl, 

Sugar, 


Faint trace 


Indican, 


Trace. 


Acetone, < 


Strong trace. 


Bile, 


Moderate amount. 


Blood, 

€tC. 


Large amount. 
Very large amount. 




^ Excessively large amount 


For Sediments use: — 


Occasional. 


Few. 


Moderate number. 


Many. 


Very many. 


Excessivel 


y large number. 



1 After Judson Daland. 



284 APPENDIX. 

OFFICIAL REQUEST FOR WIDAL REACTION, COUNTY AND 
CITY OF PHILADELPHIA, PENNA. 

Page 1 

TO BE FILLED OUT BY ATTENDING PHYSICIAN. 

Is this the first specimen from this case ? Date 

Name of Attending Physician 

Residence 

Name of Patient Age 

Residence Ward 

All data must be given or report will not be made. 
(See inside for Directions.) 



Page 2 

TO BE FILLED IN BY ATTENDING PHYSICIAN. 



Day of fever Has patient previously had Typhoid ? . . 

When? Sex/. 

Mark any of the following symptoms observed : 

Diarrhoea? Temp. Range Enlarged Spleen?. 

Delirium ? Iliac Tenderness ? Rose Spots ? . . 

State suspected source of infection 

Clinical Diagnosis and Remarks : 



Page 3 

DIRECTIONS FOR TAKING BLOOD DROP. 

Thoroughly cleanse the skin of the patient's finger tip or lobe of 
the ear. After carefully drying, prick it with a needle previously 
sterilized by heating over a lamp or gas flame and allowed to cool. Allow 
five or more large drops of blood to dry on the inner surface of the 
accompanying piece of folded paper. Fill out the first two pages of 
this book. Replace the folded paper and book in the colored envelope, 
which should be stamped and posted or otherwise returned to the Labora- 
tory. Room 708, Citv Hall. 



Page 4 

CIRCUMSTANCES INFLUENCING THE INTERPRETATION OF 
THE RESULTS OBTAINED. 

The test is not likely to yield positive results before the fifth or 
sixth day of an attack of typhoid fever, but the best results are obtained 
about the tenth day. 

The condition of the blood causing the reaction persists for some 
time after convalescence, hence care must be taken to exclude the co- 
incidence of a positive result due to a previous attack of typhoid fever. 

In doubtful cases giving negative reaction, send further samples at 
intervals of two or three days, so long as there is a possibility of typhoid 
fever. 

Please inform the Laboratory, btf reply postal sent you, of any dis- 
crepancy between result of test and the subsequent course of case. 



APPARATUS. 



285 



APPARATUS. 

The following apparatus will be found necessary to con- 
duct the examinations described in this book : — 



FOR URINALYSIS. 



Purette and pinch-cock. 

Pipette, 1 c.c. 

Pipette, 5 c.c. 

Pipette, 10 c.c. 

Flask, 1000 c.c. 

Flask, 100 c.c. 

Graduated cylinder, 50 c.c. 

Urinometer and jar. 

Ureometer, Doremus. 

Albuminometer, Esbach. 

Plain glass pipettes. 

Glass stirring-rod. 

Porcelain dishes. 

Porcelain tile. 

Beakers. 

FOR BLOOD AND 

Gower's, FleichPs or Sahli's 
hemoglobinometer. 

Thoma-Zeiss hemocytometer. 

Drop-bottles and droppers. 

Hypodermic syringe. 

Faught or Stanton sphygmo- 
manometer. 

Daland lancet. 

Microscope and accessories. 



Sedimentation glasses. 

Covers for glasses. 

Funnels. 

Test-tubes, assorted sizes. 

Centrifuge. 

Plain centrifuge tubes. 

Graduated Purdy tubes. 

Bunsen burner and hose. 

Tripod. 

Wire gauze. 

Burette stand. 

Filter paper. 

Microscope. 

Westphal Balance. 

BLOOD-PRESSURE. 

Microscope slides. 

Cover-glass, % in. diameter. 

Mechanical stage. 

Spectroscope. 

Wright's blood-coagulation ap- 
paratus or Boggfs modifica- 
tion of Eussell and Brodies' 
apparatus. 



FOR STOMACH EXAMINATION. 

Two Mohr's burettes (2.5 and 50 cubic centimeters) will be 

required. 
Dimethyl-amido-azobenzol paper. 
Dimethyl-amido-azobenzol (Topfer's reagent). 



286 APPENDIX. 

Deci-normal sodium hydrate. 

Phenol-phthalein (1 per cent, alcoholic). 

Sol. neutral ferric chlorid. 

Congo-red paper. 

Starch paper. 

Powdered pepsin. 

Sodium carbonate. 

Potassium hydrate, 10 per cent. sol. 

Egg albumin in cubes or buttons preserved in glycerin. 

Mett^s Capillary Albumin Tubes. 



FOR FECES, CEREBRO-SPINAL FLUID AND MILK. 

Stool sieve. 
Breast-pump. 

Hydrometer, 1010 to 10-10. 
Graduated cream-gauge. 
Cream centrifuge tube and pipettes. 

Capillary tube, heavy glass. Diameter, 0.05 mm. Length, 800 
to 1000 mm. 



FOR BACTERIOLOGIC AND OPSONIC WORK. 

Incubator. McFarland & I/Engle's neph- 
1 pr. balances accurate to 0.2 elometer. 

gram. Arnold steam sterilizer. 

Bunsen burner. Thermo-regulator. 

Thermometer registering 200° Eubber tubing. 

C. Platinum wire loops, glass 
Thermometer graduating in handle. 

tenths. Eegistering from Sterile bouillon in tubes. 

to 50° C. Agar slants. 

Large watch-crystals. Capillary pipettes. 

Pure culture of typhoid bacil- Hanging drop slides. 

lus. 



CHEMICALS AND REAGENTS. 



287 



CHEMICALS AND REAGENTS. 

The following chemicals and reagents will be required to 
carry out the examinations outlined in this work: — 



CHEMICALS-FOR URINALYSIS. 



Acid, nitric. 

sulphuric. 

hydrochloric C. P. 

hydrochloric deci-normal. 

acetic, glacial. 

acetic, 10 per cent. 

salicylic. 
Ammonium hydrate. 
Potassium hydrate (sticks). 

" 20 per cent, solution. 
Sodium hydrate (sticks). 

" 40 per cent, solution. 

" deci-normal solution. 
Silver nitrate, standard sol. 
Copper sulphate, 1 to 10 sol. 
Ammonium sulphate, sat. sol. 
Sodium chlorid, sat. sol. 
Litmus papers. 
Ethyl alcohol. 
Methyl alcohol. 
Amylic alcohol. 
Potassium chlorate, 1 per cent. 

sol. 
Lead acetate (trebasic). 



Sodium acetate. 

Potassium ferrocyanid. 

Zinc chlorid, 5 per cent. aqu. 
sol. 

Potassium acetate. 

Chloral. 

Sodium carbonate. 

Neutral ferric chlorid. 

Bajium chlorid, standard sol. 

Sodium nitroprussicl. 

Solution hydrogen dioxicl. 

Bromine water. 

Phenylhydrazin hydrochlorid. 

Ethylene diamine hydate, 10 
per cent, aqueous. 

Zinc acetate, 10 per cent, al- 
coholic sol. 

Phenolphthalein, 1 per cent. 
alcohol, sol. 

Powdered guaiac. 

Orcin. 

Chloroform. 

Lead carbonate. 



Normal saline solution. 

Benzol. 

Xylol. 

Chloroform. 

Litmus paper, red and blue. 

Canada balsam. 



FOR BLOOD. 

Xylol balsam. 
Xylol dammar. 
Ether. 

y 2 per cent, acetic acid. 
2 1 /2 per cent, potassium bi- 
chromate. 



288 APPENDIX. 

REAGENTS— FOR URINE. 

Magnesia Mixture: — 

Ammonium chlorid 1 part. 

Magnesium sulphate 1 part. 

Ammonia water 1 part. 

Water 8 parts. 

The salts are dissolved in the water and the ammonia 
water then added. 

Knop's Solution: — 

Bottle "A"— Sodium hydrate sol 1 : 25 

Bottle "B"— Bromine 1 part. 

KBr 1 part. 

Water 8 parts. 

For the test add 1 part of the bromine solution to 15 
or 20 parts of the sodium hydrate solution. 

Purdy's Keagent: — 

Potassium ferrocyanid 10 parts. 

Strong acetic acid 10 parts. 

Water 10 parts. 

Tanret's Keagent: — 

Dissolve 33.1 grams potassium iodid in 200 cubic cen- 
timeters of water. Add 13.5 grams powdered mercuric 
chlorid and water, stirring until the red precipitate first 
formed has been dissolved. 

Dilute to 900 cubic centimeters with water and add 
100 cubic centimeters strong acetic acid. Allow to stand 
twelve hours, and then decant from precipitate and use 
clear solution. This forms a solution of mercuric-potassium 
iodid in dilute acetic acid. 

Esbach's Eeagent: — 

Picric acid 10 grams. 

Citric acid 20 grams. 

Water 1000 c.c. 



CHEMICALS AM) REAGENTS. 2S ( .) 

Feiilixg's Reagent: — 

The reagent consists of two solutions which are kept 
in separate bottles until mixed immediately before using. 

Solution "A" — Copper sulphate 34. 64 grams. 

Water 500.00 c.c. 

Solution "B" — Sodium-potassium tartrate 173.00 grams. 

Sodium hydroxid 125.00 grams. 

Water 500.00 c.c. 

These solutions are used in equal parts for the test. 

Xylaxder's Reagent : — 

Bismuth subnitrate 2 grams. 

Rochelle salt 4 grams. 

Sodium hydroxid (8 per cent, sol.) 100 c.c. 

Trommer-Simrock Reagent : — 

Copper sulphate 2 grams. 

Potassium hydroxid (5 per cent. sol.). . . . 150 c.c. 

Glycerin 15 c.c. 

Distilled water 15 c.c. 

Purdy's Reagent for Sugar: — 

Copper sulphate 4.72 grams. 

Glycerin 38.00 c.c 

Water 200.00 c.c. 

These should be dissolved in the water by gentle heat. 

Potassium hydroxid 23.50 grams. 

Water 200.00 c.c. 

Dissolve separately and then add to the copper solution. 
When cold add: — 

Ammonium hydroxid (strong) 450.00 c.c. 

Water, q. s 1000.00 c.c. 

Diazo Reagent: — 

This reagent consists of two solutions which are kept 
separate until mixed for the test. 

Solution "A" — Sulphanilic acid 1.0 gram. 

Hydrochloric acid (con.) . . . 50.0 c.c. 

Water 1000.0 c.c. 

Solution "B" — Sodium nitrite 1.0 c.c. 

Water . 200.0 c.c. 

Proportion for test : "A," 5 c.c. "B," 3 drops. 

19 



290 APPENDIX. 

FOR BLOOD. 

Hayem's Diluting Solution for Counting Eed Blood- 
Cells : — 

Mercuric bichlorid 0.5 gram. 

Sodium sulphate 5.0 grams. 

Sodium chlorid 1.0 gram. 

Aq. dest 200.0 c.c. 

Toisson's Solution for Simultaneously Counting Eed and 
White Cells: — 

Methyl violet 0.05 gram. 

Neutral glycerin 30.00 c.c. 

Aq. dest 80.00 c.c. 

Mix and add : — 

Sodium chlorid 1.00 gram. 

Sodium sulphate 8.00 gram. 

Aq. dest 80.00 gram. 

Filter. Twelve minutes required to stain white blood- 
cells. 

Fixing Methods: — 

1. Equal parts of absolute alcohol and ether. The 
specimen should be immersed in this for half to two hours. 
This method is particularly good for malarial parasites and 
degeneration in blood-cells. 

2. Absolute alcohol for five minutes. 

3. Flemming's solution: — 

Chromic acid, 1 per cent 15 parts. 

Osmic acid, 2 per cent 4 parts. 

Glacial acetic acid 1 part. 

The blood-specimens, as soon as they are made, before they 
have time to air-dry, are plunged into this solution and allowed 
to remain for ten minutes. They are then washed in running 
water for ten minutes and dried. This solution is very useful 
for demonstrating the chromatin of nuclei. 

4. Vapors of formaldehyde. 

5. Heat. 



CHEMICALS AND REAGENTS. 291 

6. Wood alcohol. This is coining rapidly in favor, as it 
can be mixed with the stain, thus reducing the time of preparing 
specimens. 

FOR GASTRIC ANALYSIS. 

Uffleman's Keagent: — 

Carbolic acid (4 per cent.) 10 c.c. 

Water 20 c.c. 

Liquor f erri chloridi 1 drop. 

This solution should be a clear amethyst color, and should 
be prepared fresh for use. 

Lugol's Iodin Solution: — 

Iodin 1 part. 

Potassium iodid 2 parts. 

Water 50 parts. 

Gunzberg's Phloroglucin Vanillin: — 

Phloroglucin 2 gms. 

Vanillin 1 gm. 

Alcohol 30 c.c. 

This solution, if active, is pale yellow. It darkens and 
deteriorates with age, especially on exposure to light, so should 
be kept in colored bottles and made fresh from time to time. 



FOR CEREBROSPINAL FLUID AND MILK. 

Saturated aqueous solution of metlryl violet. (E 5.) 

Cream Testixg Solutions : — 

"A" — Amylic alcohol 37 parts by volume. 

Methyl alcohol 13 parts by volume. 

Hydrochloric acid 50 parts by volume. 

"W— Sulphuric acid. sp. gr. 1832. 



292 APPENDIX. 

STAINS. 

Ehrlich's "Triacid" Staix: — 

Orange G 13.0-14.0 c.c. 

Acid fuchsin 6.0- 7.0 c.c. 

Distilled water 15.0 c.c. 

Methyl green 25.5 c.c. 

Alcohol 10.0 c.c. 

Glycerin 10.0 c.c. 

The three stains, orange G, acid fuchsin, and methyl green, 
are prepared in saturated aqueous solutions, and then mixed in 
the above amounts while being shaken thoroughly. 

Eosix axd Methylexe-Bltje : — 

Eosin 0.5 per cent, in 70 per cent, alcohol. 

Stain for a few minutes, wash, blot, and apply. 
Methylene-blue 1 per cent, aqueous. 

Stain for a few minutes. 

Chezixsky Staix: — 

Methylene-blue, sat. aq. sol 40 c.c. 

Eosin, 0.5 per cent, in 70 per cent, alcohol. ... 20 c.c. 
Distilled water 40 c.c. 

The specimens are fixed in absolute alcohol for from five 
to thirty minutes, and stained in a thermostat at 37° C. for 
from three to six hours. 

Hematoxylix-Eosix (Ehrlich's mixture) : — 

Eosin (crystals) 0.5 gram. 

Hematoxylin 2.0 gram. 

Absolute alcohol, 1 

Distilled water, i of each 100.0 gram. 

Glycerin, 

Glacial acetic acid 10.0 gram. 

Alum in excess. 

This mixture should be allowed to stand for several weeks 
before it is ready for use. The specimens are stained in from 
half to two hours. 

Carbol Gextiax-Yiolet : — 

Gentian-violet, cone, alcoholic sol 10.0 c.c. 

Carbolic acid, 5 per cent, watery sol 100.0 c.c. 



STAINS. 293 

Giemsa Improved Stain: — 

Azur 11 eosio 3.0 gram. 

Azur 11 0.8 gram. 

Exsiccate, pulverize, and sift. 
Dissolve in chemically pure glycerin at 

60° C 250.0 c.c. . 

When solution is complete add : — 
Methyl alcohol at temperature 60° C 250.0 c.c. 

Shake well, allow to stand for twenty-four hours, then 
filter. 

Decolorizing Solutions : — 

I. Acetic acid, 0.5 to 5.0 per cent, watery solution. 
II. Xitric acid, 20 to 30 per cent. 
III. Acid alcohol: — 

Sulphuric acid (cone.) 30 drops. 

Alcohol (95 per cent.) 50 c.c. 

Water 150 c.c. 

Loeffler's Alkaline Methylene-Blue : — 

Concent, alcohol solution methylene-blue . . . 30.0 c.c. 
Potassium hydrate (%oooo) 100.0 c.c. 

Ziehl's Carbol-Fuchsin : — 

Fuchsin in substance 1 gm. 

Carbolic acid (cryst.) 5 gm. 

Alcohol (95 per cent.) 10. c.c. 

Distilled water 100. c.c. 

Or it may be prepared by adding to a 5 per cent, watery 
solution of carbolic acid a saturated alcoholic solution of 
fuchsin until a metallic luster appears on the surface of the 
liquid. 

Gabbott's Method: — 

A — Fuchsin 1 gm. 

Absolute alcohol . 10 c.c. 

Carbolic acid (5 per cent.) . . : 100 c.c. 

B — Methylene-blue 2 gm. 

Sulphuric acid (35 per cent, sol.) 100 c.c. 



294 APPENDIX. 

Gram's Iodine: — 

Iodin 1 gm. 

Potassium iodid 2 gm. 

Distilled water 300 c.c. 

KoCH-ElIRLICH GENTIAN-VlOLET : 

Take distilled water, 100. cubic centimeters, and add 
anilin oil, drop by drop, until the solution has an opalescent 
appearance. The vessel containing the mixture should be 
thoroughly shaken after the addition of each drop. It is then 
filtered through moistened filter paper until the filtrate is clear. 
To 100 cubic centimeters of the filtrate add 10 cubic centi- 
meters of absolute alcohol and 2 cubic centimeters of concen- 
trated solution of gentian-violet. 

Unna's Orcein Stain: — 

Orcein in substance 1 gm. 

Hydrochloric acid 1 c.c. 

Absolute alcohol 100 c.c. 

Eeagents for Staining Flagella : — 
Mordant. 

Tannic acid (20 ac. to 80 water) 10 c.c. 

Ferric sulphate cold sat. sol 5 c.c. 

Fuchsin sat. watery sol 1 c.c. 

Adjuvants. 

Sodium hydrate 1 per cent, aqueous solution. 

Sulphuric acid (1 c.c. equal 1 c.c. of 1 per cent. NaOH). 

Pappenheim's Solution: — 

Corallin 1 part. 

Absolute alcohol 100 parts. 

Add to the above solution methylene-blue in bulk to sat- 
uration. Finally add 20 parts of glycerin. 

Loeffler's Mordant : — 

Tannic acid solution (2 parts acid, 80 parts 

water) 10. c.c. 

Ferrous sulphate saturated solution 5. c.c. 

Fuchsin saturated aqueous solution 1. c.c. 



STAINS. 295 

Ebner's Fluid : — 

Hydrochloric acid 2.5 c.cm. 

Sodium chlorid, C. P 2.5 c.cm. 

Distilled water 100.0 c.cm. 

Alcohol, 95 per cent 500.0 c.cm. 

Universal Staining Method: — 

The discovery of the process whereby two or more colors 
could be chemically combined in one staining reagent, marked 
a great advance in the field of hematology. It affords greater 
opportunity for more detailed study of the minute structure of 
the cells of the blood, which in turn has resulted in a better 
classification of the different elements through the differentia- 
tion of new and distinct varieties which, until recently, have 
been unrecognized. Of these combination stains the Polychrome 
or "universal" staining method is by far the best and most prac- 
tical, and is now rapidly superseding the older and more cum- 
bersome methods. For this reason it becomes a matter of con- 
siderable importance, almost a necessity, that the clinical worker 
should have at hand one or more of these stains ready for imme- 
diate use when occasion requires. 

Unfortunately the preparation of this class of stain in- 
volves considerable expenditure of time, and demands no small 
amount of chemical knowledge and manipulative skill. These 
factors combine to limit the preparation of these stains to a 
comparatively small number of experienced workers, while the 
majority of the profession is dependent upon unstable liquid 
stains, obtainable through the supply houses, the composition 
of which is frequently so variable as to render the results value- 
less. Further, all aniline stains of this character are prone to 
decomposition w 7 hen kept even for a moderately long time in 
solution. 

Fortunately the demand for a uniform and reliable stain 
has recently been met by a London Pharmaceutical House, 2 who 
now carry in stock a number of very uniform and perfectly re- 
liable stains under the name of "Soloid" brand. These stains 
are very carefully prepared and their composition practically 



2 Burroughs Wellcome & Co., with a branch at 45 Lafayette Street, New York City. 



296 



APPENDIX. 



uniform. A definite quantity of the dried stain is compressed 
into a tiny tablet and dispensed in vials containing six. Each 
package is accompanied with information indicating the proper 
dilution and best working conditions for that particular stain. 

The "Soloid" stains permit of the preparation of small 
quantities of liquid stain whereby waste through evaporation or 
decomposition is reduced to a minimum, at the same time the 
results obtained, as far as the author has employed them, are in 
every way satisfactory. 

The following is abstracted from the last descriptive cata- 
logue of the above company: — 

The majority of the stains are employed in alcohol in solu- 
tion, and the different alcohols commonly used are here described. 

Absolute Alcohol contains not less than 99 per cent, by 
weight of pure ethyl alcohol, C 2 H 5 OH. 

Alcohol of a Stated Percentage, e.g., 50 per cent, alco- 
hol, means a mixture with water wmich contains the stated per- 
centage, i.e., 50 per cent, by volume of pure ethyl alcohol. 

Methyl Alcohol is a pure substance, CH 3 OH, prepared 
by the purification of commercial wood spirit. Commercial 
methyl alcohol, which is impure, must not be employed in the 
preparation of Jenner's, Leishmam's, or Eomanowski stains. . 

The amounts of distilled water and absolute alcohol re- 
spectively, required to produce saturated solution of certain dyes 
in common use, are indicated in the following table : — 





4 'Soloid" 
product of 

0.1 grm. 

of dye 


Water 
(c.c.) 


Alcohol 
(c.c.) 


Bismarck Brown, pure 


1 
1 
1 
1 
1 
1 
1 


7 
10 

7 
2 
5 
7 
5 


7 


Fuchsin, " 


2.5 


Gentian-Violet, s * 


7 


Hematoxylin, ' 


1 


Methyl Violet, " 


1 


Methylene-Blue, ic 


7 


Thionin Blue, " 


10 



Aqueous dilutions of the above, containing 5 to 10 per 
cent, of these saturated solutions, are well adapted for ordinary 
staining purposes. Various other solutions, ready for imme- 
diate use, may be prepared from "Soloid" Microscopic Stains 
according to the following directions: — 



STAINS. 297 

Eosix. — To obtain a solution of eosin suitable for general 
staining, one "Soloid" product may be dissolved in &0 cubic 
centimeters of 50 per cent, alcohol. This gives a O.o per cent. 
solution. 

Loeffler's Alkaline Methylene-Blue. — Dissolve one 
"Soloid" methvlene-blue in 7 cubic centimeters of absolute 
alcohol, and add 25 cubic centimeters of distilled water, to 
which one drop of Liquor Potassae U.S. P. has been added. 

Aniline Gentian-Violet. — Dissolve one "Soloid" gen- 
tian-violet in T cubic centimeters of absolute alcohol, and add 
63 cubic centimeters of a freshly filtered saturated solution of 
aniline oil in distilled water. 

Carbol Gentian- Violet. — Dissolve one "Soloid" gentian- 
violet in 7 cubic centimeters of absolute alcohol, and add 63 
cubic centimeters of a 1 per cent, aqueous solution of carbolic 
acid. 

Ziehi/s Carbol-Fuchsin. — Thoroughly powder and dis- 
solve one "Soloid" fuchsin in 3 cubic centimeters of absolute 
alcohol, add 22 cubic centimeters of 5 per cent, aqueous solu- 
tion of carbolic acid; shake well, and filter before using. 

Gram's Iodine Solution. — Dissolve one "Soloid" product 
of reagent A in 10 cubic centimeters of distilled water, add 
one of reagent B, and when solution is complete, dilute to 15 
cubic centimeters with distilled water. 

Carbol Thionin Blue. — Dissolve one "Soloid" thionin 
blue in 100 cubic centimeters of a 5 per cent, aqueous solution 
of carbolic acid. 

Borax Methylene-Blue. — Dissolve one "Soloid" borax 
methylene-blue in 10 cubic centimeters of distilled water. 

Delafield's Hematoxylin. — Dissolve one "Soloid" hema- 
toxylin (Delafield) in 10 cubic centimeters of a 25 per cent, 
solution of glycerin in water. 

Eosin-Azur (for Giemsa staining with one solution). — 
Dissolve one "Soloid" product in 5 cubic centimeters of pure 
methyl alcohol. 

Eosin-Methylene-Blue (Louis Jenner's Stain). — Dis- 
solve one "Soloid" product in 10 cubic centimeters of pure 
methyl alcohol. 



298 APPENDIX. 

Komanowsky Staix (Leishman's Modification). — Dissolve 
one "Soloid" product in 10 cubic centimeters of pure methyl 
alcohol. 

Sodium Carbonate. — When employed in the preparation 
of Komanowsky stain, dissolve one "Soloid" product (0.05 
gram) in 10 cubic centimeters of distilled water, and add one 
"Soloid" methylene-blue, 0.1 gram (for the method of prepa- 
ration and use, see British Medical Journal, September 21, 
1901, page 757). 

Biondi-Ehrlich-Heidenhain Triple Staix. — Dissolve 
one "Soloid" Ehrlich triple stain in 25 cubic centimeters of 
distilled water; one "Soloid" acid fuschin in 2 cubic centi- 
meters of distilled water, and mix. The mixture is ready for 
use and keeps well. 

Toisox Blood Fluid. — For the preservation of blood cor- 
puscles and the counting of the same. Dissolve one "Soloid" 
product in 3 cubic centimeters of glycerin and 16 cubic centi- 
meters of distilled water. The solution should always be 
filtered immediately before use. 

The Polychrome Methylexe-Blue-Eosix Staixs (Bo- 
maxowski). — There are about fifteen different modifications of 
this stain. The majority of them are difficult of manufacture, 
even by an expert, so- it is recommended that they be bought 
ready-made from the laboratory apparatus and supply houses 
which make them. 

For those who desire to make this stain for themselves, the 
following modification by Hastings is appended, as one which 
is comparatively simple of manufacture. 3 

All the Romanowski stains are made with wood alcohol, 
which, during the first portion of its application, acts as a 
fixative. 

Hastings'- Stain. — The dry stains necessary are eosin 
(water solution) yellow (Grubler), and methylene-blue (Ehr- 
lichias rectif.) (Grubler). 



3 Hastings : Johns Hopkins Hospital Bulletin, 1905. 



STAINS. 299 

Solution "A" — Eosin 1 per cent, aqueous. 

Solution "B" — Alkaline methylene-blue 1 per cent, 
aqueous. 

Solution "C" — Methylene-blue 1 per cent, aqueous. 

Solution "A" may be kept ready-made, solutions "B" and 
**("* must be made fresh. 

To prepare solution "B/ v use a warm 1 per cent, solution 
of dry powdered sodium carbonate. Add to it one per cent, of 
methylene-blue powder, and heat over a water bath for 15 
minutes. Add 30 cubic centimeters of water for each 100 cubic 
centimeters of the original fluid, and heat again for 15 minutes. 
Then pour off the solution from the residue and divide into 
two equal parts. To one part add enough 12.5 per cent, acetic 
acid solution to make a faintly acid reaction. This is best 
determined by taking a piece of blue litmus paper and allowing 
a drop to fall upon it, taking as the end reaction the point at 
which the margin of the drop after absorption in the paper 
shows a faint pink. Then add the remaining unneutralized por- 
tion to this. 

To mix the stain use distilled water 1000 cubic centimeters. 
Solution "A" 100 cubic centimeters; solution "B" 200 cubic 
centimeters: solution "C," 70 to 80 cubic centimeters. In add- 
ing solution "C" put in 70 cubic centimeters at once, and stir 
well ; if no precipitate appears, add 1 cubic centimeter at a time 
until one does appear. After the precipitate appears the stain 
is allowed to stand for half an hour and then filtered through 
one filter. Forced filtration is generally necessary. 

The dry residue is removed from the paper and pulverized. 
It may be kept in this form or dissolved in Merck's pure methyl 
alcohol. Seven- to nine-tenths of a gram of dried stain is 
usually obtained. Three-tenths of a gram is dissolved in 100 
cubic centimeters of alcohol for the staining solution. In dis- 
solving the stain it must be rubbed up with the alcohol in a 
mortar, as* the powder is with difficulty soluble. 

If there are more than nine-tenths of a gram of the dried 
stain obtained, the preparation is useless, and should be begun 
again. 



300 APPENDIX. 

For each new lot of stain made up, one must determine 
the relative proportion of stain and water used in staining, and 
the relative lengths of time during which the pure and diluted 
stain is allowed to act. 

Usually two drops of stain on the smear for one minute 
and then four drops of water added and allowed to act for four 
minutes gives the best result. 

For uniformity in dropping a dropper should be used. 

All polychrome methylene-blue stains require experiment, 
since different mixtures by the same method require slight 
variations in their use. These must be ascertained by trial. 
Use distilled water to wash the specimen, since tap-water may 
ruin it. 



TABLES. 



301 



Multiples of a drain 
From 1 grain to 1 ounce 



U. S. A. 



Metric 



gr. 1 0.065 gm. 

gr. U 0.086 gm. 

gr. 11 0.097 gm. 

gr. 1| 0.113 gm. 

gr. 2 0.13 gm. 

gr. 2\ 0.162 gm. 

gr. 3 0.194 gm. 

gr. 31 0.227 gm. 

gr. 4 0.259 gm. 

gr. 5 0.324 gm. 

gr. 6 0.389 gm. 

gr. 7 0.454 gm. 

gr. 8 0.518 gm. 

gr. 8| 0.567 gm. 

gr. 9 0.583 gm. 

gr. 10 0.648 gm. 

gr. 12 0.778 gm. 



U. S. A. 



Metric 



gr. 
gr. 
gr- 
gr- 
gr- 
gr. 
gr- 
gr. 
gr- 
oz. 
oz. 
oz. 
dr. 
oz. 
dr. 



15 0.972 gm. 

18 1.166 gm. 

20 1.296 gm. 

25 1.620 gm. 

30 1.944 gm. 

35 2.268 gm. 



40 

50 

60 

120 



2.592 gm. 
3.24 gm. 



3.89 

7.78 

i 3.54 

i 7.08 

i 14.17 

4 15.55 

1 28.35 

8 31.1 



gm. 
gm. 
gm. 
gm. 
gm. 
gm. 
gm. 
gm. 



Equivalents of U. S. A. and Metric Measures of Capacity 
From half-a-minim to 1 fluid ounce 



U. S. A. 



Metric 



min. h 0.03 c.c. 

min. 1 0.062 c.c. 

min. 2 0.123 c.c. 

min. 3 0.185 c.c. 

min. 4 0.246 c.c. 

min. 5 0.308 c.c. 

min. 6 0.370 c.c. 

min. 7 0.431 c.c. 

min. 8 0.493 c.c. 

min. 9 0.554 c.c. 

min. 10 0.616 c.c. 

min. 12 739 c.c. 

min. 15 924 c.c. 



U.S.A. 

min. 
min. 
min. 
min. 
min. 
min. 
min. 
min. 
min. 
min. 
min. 
min. 
min. 



20. 

25. 

30. 

35. 

40. 

50. 

60. 

90. 
120. 
180. 
240. 
360. 
480. 



Metric 


1.232 


c.c 


1.54 


c.c 


1.848 


c.c 


2.156 


c.c 


2.464 


c.c 


3.08 


c.c 


3.70 


c.c 


5.54 


• C 


7.39 


c.c 


11.09 


c.c 


14.79 


c.c 


22.18 


c.c 


29.57 


c.c 



In Continental prescribing, a smaller quantity than half a cubic centimeter is 
usually expressed in drops, which, in dispensing:, are dropped from pipette into the 
cubic centimeter measure. 



302 



APPENDIX. 



Approximate U. S. A. Equivalents of Metric Measure of Capacity 



Metric 


U. S. A. 


Metric 










U. S. A. 


1 c.c 


16 (16.23) min. 

32 i min. 

48} min. 


25 c.c. 






6fl 


Hr 


, 46 min. 


2 c.c 


30 c.c. 
40 c.c. 






. 8fl. 
,2fl. 


dr. 
dr. 


, 7 min. 


3 c.c 


. lfl. 


oz. 


49 min. 


4 c.c 


1 fl. dr. 5 min. 


50 c.c. 


. lfl. 


oz. 


, 5fl. 


dr. 


, 32 min. 


5 c.c 


lfl. dr. 21 min. 


75 c.c. 


. 2fl. 


oz. 


, 4fl. 


dr. 


17 min. 


6 c.c 


lfl. dr. 37 min. 


100 c.c. 


. 3fl. 


oz. 


3fl. 


dr. 3 


3 min. 


7 c.c 


lfl. dr. 54 min. 


125 c.c. 


. 4fl. 


oz. 


lfl. 


dr. 


49 min. 


8 c.c 


2fl. dr. 10 min. 


150 c.c. 


. 5fl. 


oz. 


Ofl. 


dr., 


35 min. 


9c. c 


2fl. dr. 26 min. 


200 c.c. 


. 6fl. 


oz. 


6fl. 


dr., 


6 min. 


10 c.c 


2fl. dr. 42 min. 


300 c.c. 


.10 fl. 


oz. 


, lfl. 


dr. 


9 min. 


12 c.c 


3fl. dr. 23 min. 


500 c.c. 


.16 fl. 


oz. 


7fl. 


dr., 


15 min. 


15 c.c 


4 fl. dr. 4 min. 


1 litre. . 


.33 fl. 


oz. 


, 6fl. 


dr. 


, 31 min. 


20 c.c 


5fl. dr. 25 min. 















Approximate U. S. A. Equivalents of Metric Measures of Mass 



Metric 



U. S. A. 



1 mgm eV gr« 

2 mgm 3 X 3 gr. 

3 mgm 2T gr 

4 mgm 3-V gr 

5 mgm T J 3 gr. 

6.5 mgm T \ gr. 

8 mgm J gr. 



Metric 



U.S.A. 



* gr. 
t gr« 
i gr. 
f gr. 



1 cgm 

2 cgm 

3 cgm 

5 cgm 

6.5 cgm 1 gr, 

10 cgm 1J gr, 

15 cgm 2J gr 

20 cgm 3 gr, 

26 cgm 4 gr 

30 cgm 4 J gr 

40 cgm 6\ gr 

50 cgm 7| gr, 

75 cgm 11 J gr. 



1 gm 15^(15.432) gr. 

2 gm 30 J gr. 

3 gm 46^ gr. 

4 gm 61| gr. 

5 gm 77£-gr. 

7.5 gm 115|gr. 

10 gm 154£ gr. 

15 gm 231igr. 

20 gm 308|gr. 

25 gm 385igr. 

30 gm 1 oz. 25 J gr. 

40 gm 1 oz. 179f gr. 

50 gm 1 oz. 334 gr. 

75 gm 2oz. 282Jgr. 

100 gm 3 oz. 230| gr. 

150 gm 5oz. 127jgr. 

250 gm 8oz. 358 gr. 

500 gm lib. loz. 278 gr. 

750 gm lib. 10 oz. 200 gr. 

1 kgm 21b. 3 oz. 120 gr. 



INDEX 



Abbe's condenser, 4 
Abnormal constituents of urine, 194 
Acetic acid-ether-guaiac test, 132 
Acetic acid in stomach, 130 
Acetonuria, 212 

ethylene-diaminhydrate test, 213 

Legal's test, 213 
Acids of digestion, 126 
Actinomycosis, pulmonary, 30 
Agar-agar, 266 
Albumin in urine, 194 

albumosuria, 198 

Bence Jones, 399 

fibrin, 200 

nucleo-albumin, 200 

proteose, 200 

qualitative tests for, 196 

quantitative tests for, 198 

serum-albumin test, 200 
-globulin test, 200 

transient albuminurias, 194 
Albumosuria, 198 

test for, 199 
Alcohol in stomach, 130 
Alkaline urine, 173 
Alkaptonuria, 165 
Ameba coli, 103 

characteristics of, 103 
examination for, 104 

dysenterise in feces, 157 
Anemias, the, 52 

chlorosis, 54 

primary, 53 

progressive pernicious, 53 

secondary, 52 
Angina pectoris, 86 

sphygmomanometer in, 87 
Anguillula intestinalis et stercoralis, 113 
Animal parasites, 101 

classification of, 101-102 

temporary, 101 

permanent, 101 
Ankylostoma duodenale, 115 
Anuria, 167 
Apparatus for bacteriologic work, 299 

for blood work, 288 

for opsonic work, 299 

for urinalysis, 288 
Appendix, 274 
Arnold sterilizer, 261 
Ascaris lumbricoides, 116 

trichiura, 114 
Autoclave sterilization, 259-261 



B 



Bacillus coli communis, 154 
morphology of, 154 
in feces, 154 
dysentericus, 156 
lactis aerogenes. 155 
of. diphtheria, 250 
of influenza, 249 
of Pfeiffer, 249 
proteus vulgaris, 155 



Bacillus, tuberculosis, 244 
typosis, 155 

morphology of, 155 
Bacteria in urine, 16 
Bacteriologic methods, 244 
blood serum in, 267 
diplococcus pneumonii, 248 
gonococcus of Neisser, 251 
Gram's stain, 252 
incubator for, 271 
Loeffler's method, 254 
Petri dish method, 269 
plate method, 270 
preparation of media, 263 
staining flagella, 254 
sterilization, 255 
Wright's stain, 252 
Balantidium coli, 107 
Bile acids in the urine, 215 
pigments in the feces, 157 
in the urine, 214 
Gmellin's test for, 214 
significance of, 214 
Bird's formula, 172 
Body fluids, 236 
Bogg's coagulometer, 93 
Bothriocephaloidiae, 109 

bothriocephalis latus, 109 
Bottger's bismuth test, 203 
Bouillon, 263 
Blood, 31 
agglutination reaction, 61 
appearance of, 32 
bacteriologic examination of, 58 
-cells, 15 

chemical composition of, 31 
color of, 33 
color-index of, 40 
-count, 49 
differential, 49 
normal differential, 50 
counting red cells, 37 
cultures, value of, 58 
estimation of hemoglobin, 35 
examination form, 279 
examination of, 16 
counting red cells, 16 

white cells, 17 
stained specimen, 17 
fixing smears of, 44 
leukocytes in, 48 
odor of, 33 
parasites, 96 

percentage of red cells, 40 
-pressure 76 
arterial, 79 
diastolic, 81 

direct method of observation, 76 
effect on urine excretion, 88 
hypertension, 79, 84 
hypotension, 84 

indirect method of observing, 77 
method of recording, 83, 281 
pathologic variations of, 83 
postural variations of, 83 



(303) 



304 



INDEX. 



Blood-pressure, pulse pressure, 82 
relation to vessel-contraction, 77 
to heart-power, 77 

reaction of, 34 

report forms, 281 

-serum media, 267 
Councilman and Mallory's, 268 
Loeffler's mixture for, 278, 279 

smears, 31 
preparation of, 43 

specific gravity of, 33, 34 

sphygmomanometer in. 79 

spirochaete pallida in, 59 

staining smears of, 45, 46 

systolic pressure, 80 

taste of, 33 

tone of vessel walls, 78 

total quantity of, 34 

Widal reaction in, 61 



Camera lucida, 17 

method of using, 18 
Cammidge reaction, 209 
Capacity of stomach, 125 
Care of the microscope, 7 
Casts in the urine, 218 

diagnosis of, 13 

preparation of sediment of, 218 

search for, 13 

varieties of in, 219 
Cercomonides, 105 

cercomonas intestinalis, 105 

lamblia intestinalis, 105 
Cerebrospinal fluid. 231 

cell-content of, 232 

differential cell-count, 233 

pressure, measurement of, 231 

proteid-content of, 234 

technic of puncture, 231 

the specimen, 232 
Chemicals for urinalysis, 285 
Chlorides in the urine, 180 

estimation of, 181 
Chlorosis, 54 
Chyluria, 222 

Clinical laboratory report forms, 277, 
blood, 279 
blood-pressure, 281 
feces, 282 
gastric, 280 
sputum, 277 
urinalysis, 277 

terms, 283 

value of sphygmomanometer, 85 
Coagulation time of the blood, 89 

Bogg's coagulometer, 93 

general considerations, 89 

in typhoid fever, 94 

methods of determining, 89 

Milan's method, 89 

normal time, 94 
averages, 95 

Russell and Brodie's method, 93 

specimen for testing, 89 

Wright's method, 90 
Coccidia hominis, 106 

perforans, 106 
Color-index of blood, 40 
Comma bacillus, 156 

morphology of, 156 

cultural characteristics of, 156 
Concretions in the urine, 225 

analysis of, 226 
Creatinin, 191 
Crystals in urine, 15 



282 



Cubic centimeters to minims (table), 301 
Culture media, 263 

agar-agar, 266 

blood-serum, 267 

bouillon, 263 

gelatin, 265 

glycerin agar-agar, 266 

preparation of, 263 
Curschmann's spirals, 27 
Cylindroids, 14 

diagnosis of, 14 

microscopic appearance of, 221, 222 
Cylindruria, 222 
Cysticercus acanthotrias, 111 
Cystin in the urine, 193 

tests for, 193 



Daland-Faught gastric apparatus, 118 
Daland hematokrit, 39 
Detection of elastic fibers, 29 

of malarial parasite, 97 
Determination of alkalinity of urine, 174 

of functions of stomach, 118 

of gastric acidity, 128 

of urine acidity, 174 
Dextrose, 210 

Diabetes, sphygmomanometer in, 87 
Diacetic acid, test for, 213 
Diaphragm, iris, 4 
Diastolic blood-pressure, 81 

definition of, 82 

methods of obtaining, 81 

recording, the, 83 
Diazo reaction, 226 

significance of, 227 

test, 226 
Dicrocelium lanceolatum, 108 
Differential leukocyte-count, 49 
Dimethyl-amido-azobenzol test, 127 
Diphtheria bacillus, 250 
characteristics of, 251 
staining of, 251 

blood-pressure in, 88 
Diplococcus pneumonii, 248 

staining capsule of, 249 
Disinfection, 262 

of feces, 263 

of sputum, 263 
Distomum hepaticum, 107 

lanceolatus, 108 

pulmonale, 108 
in lung, 30 

Westermanni, 108 
Dittrich's plugs, 25 
Donne's test for pyuria, 217 
Drugs in the urine, 227, 230 

antifebrin, 229 

antipyrine. 229 

bromine, 228 

copiaba, 229 

iodine, 228 

lead, 227 

mercury, 227 

phenacetin, 229 

pyramadin, 229 

salicylic acid, 229 

santol oil, 229 

santonin, 23C 

tannic acid, 229 

E 

Ebner's fluid, 299 
Echinococcus disease of lung, 30 
Ehrlich's myelocytes, 49 
Elastic fibers, detection of, 29 



INDEX. 



305 



Eosinophilic leukocytes, 49 

Epithelia, 14 
detection of, in urine, 218 
general characteristics of, 14 

Estimation of hemoglobin, 35 
method of Fleishl, 35 
of peptic activity, 132 
of glucose in urine, 205 

Ewald test -breakfast, 118 
modified, 121 

Examination of the blood, 16 

Eye-piece or ocular, 2 
finder in, 3 



Faught sphygmomanometer, 82 

application of, 79 
Fecal concretions, 158-160 
Feces, 143 

after test-diet, 147 

albumin fermentation of, 158 

ameba dysenteriae in, 157 

bacillus coli communis in, 154 
dysentericus in, 156 
lactis aerogenes in, 155 
of Shiga in, 156 
proteus vulgaris in, 155 
typhosis in, 155 

bile pigment in, 157 

blood in, 148, 150, 153 

calculi in, 148 

carbohydrate fermentation of, 158 

color of, 144 

comma bacillus in, 156 

connective tissue in, 149, 158 

consistence of, 143 

crystals in, 149 

epithelia in, 150 

fat in, 158 

fermentation test, 152 

foreign bodies in, 159 

gall-stones in, 159 

lost-albumin test, 152 

macroscopic appearance, 149 

meat-fiber in, 158 

microscopic appearance, 149 

mucus in, 157 

muscle fiber in, 149 

normal appearance of, 148 

parasites in, 148, 157 

pathologic appearance of, 148 

physical characteristics of, 143 

potato cells in, 149 

pus in, 148, 150 

reaction of, 150 

report form for, 282 

serous stools, 143 

streptococcus aerogenes in, 155 

sublimate test, 152 
Fehling's sugar test, 201 

quantitative test, 205 
Fermentation of feces, 158 

saccharometer, 206 
Fibrin in urine, 200 
Filaria sanguinis hominis, 99, 113 

nocturna, 99 
Flagellata, 105 
Flat-worms, 107 
Free hydrochloric acid, 127 

detection of, 127 

quantitative estimation of, 129 
Fresh blood preparation, 43 

G 

Gall-stones, 159 
chemical examination of, 160 



Gastric contents, 118 

examination blank forms, 280 

lavage, 126 

microscopic examination of, 131 

test-meal removal, 122 
Gastro-intestinal tract, 145 

motor functions of, 146 

Schmidt diet, 146 
Gelatin, nutrient, 265 
Glucose in the urine, 201 

Bottger's test for, 203 

Cammidge reaction, 209 

Fehling's qualitative test, 201 
quantitative test, 205 

fermentation test for, 206 

Nylander's test for, 203 

phenyl hydrazin test, 204 

polarimetric test for, 211 

Purdy's quantitative method, 208 

Robert's differential test for, 206 

Trommer's test for, 204 
Glutoid capsule test, 144 
Glycerin agar-agar, 266 
Glycosuria, 205 

clinical significance of, 211 

temporary, 211 
Gonococcus of Neisser, 251 

cultural characteristics of, 251 

differentiation of, 252 

microscopic appearance of, 251 

staining of, 252 
Gower's hemocytometer, 35 
Grains to grams (table), 300 
Gram's method, 252 

Wright's modification of, 252 
Grams to grains (table), 301 
Gram-positive organisms, 253 

-negative organisms, 253 
Gunzberg's test, 128 

H 

Hammerschlag's test for pepsin, 133 
Heart-failure cells, 27 

-muscle degeneration, 86 
Hematuria, 215 

microscopic appearance of, 216 

test for, in urine, 216 
Hemoglobin, estimation of, 35 

method of Gower, 35 
Hemoglobinuria, 217 
Hemoptysis, 24 

causes of, 24 
Hemosporidise, 107 
Hippuric acid, 191 

microscopic appearance of, 191 
Hot air sterilization, 262 
Human milk, 240 

characteristics of, 240 

composition of, 243 

determination of fat in, 241 
of proteids in, 243 
of sugar in, 242 

examination of, 241 

quantity of, 241 

reaction of, 241 

specific gravity of, 241 

the sample, 240 
Hydrocele fluid, 239 
Hydrochloric acid in stomach, 126 

estimation of, 128 
Hydruria, 167 
Hypotonus, 79 

I 
Illumination of microscope, 6 
Increased acidity of urine, 173 



306 



INDEX. 



Incubator, 271 

therruo-regulator for, 272 
Indican in urine, 179 

test for, 180 
Inflation of stomach, 123 

contraindications to, 124 
Influenza bacillus, 249 

staining for, 250 
Infusoria, 107 
Intestinal digestion, 144 

glutoid capsules in, 144 

iodine test in, 145 

salicylic acid test in, 146 
Iodtiie reaction in saliva, 145 
Iodoform test, 138 
Iris diaphragm, 4 

K 

Kidney disease, blood-pressure in, 86 



Lactic acid in gastric-contents, 130 

test for, 130 
Lactoseuria, 212 
Lamblia intestinalis, 105 
Large mononuclear leukocytes, 48 
Lead colic, blood-pressure in, 88 
Legal's test, 213 
Leischman-Donovan bodies, 100 

staining of, 100 
Leucin, 193 
Leukemia, 54 

blood-plaques in, 55 

leukocytic, 56 

lymphocytic, 55 

myelocytes in, 56 
Leukocytes, estimation of, 42 

Thoma-Zeiss method, 42 
Leukocytosis, 51 

pathologic, 51 

physiologic, 51 
Leukopenia, 52 
Liver fluke, 107 

Loeffler's staining for flagella, 254 
Lowered gastric secretion, 129 

detection of, 129 
Lymphocytes, 48 

M 

McFarland and L'Engle's nephelometer, 73 
Malaria, Plasmodium of, 96 

clinical relations of, 98 

detection of, 97 

differentiation of (table), 97 
Maltosuria, 212 
Maltwood finder, 9 

method of using, 10 

Pepper's guide for, 11 
Mast cells, 49 
Mechanical stage. 8 

description of, 8 

Vernier scale on, 9 
Meeker's albumin test, 197 
"Melangeur," 37 
Melanuria, 165 

test for, 165 
Method of Leo, 136 
Mett's pepsin method, 133 
Metz's formula, 172 
Microscope, 1 

barrel of, 3 

care of, 1, 7 

continental type of, 5 

draw tube of, 4 

English type of, 5 



Microscope, fine adjustment of, 4 

illumination of, 6 

nose piece of, 5 

oblique illumination of, 6 

reflectors of, 4 

selection of, 1 

stage of, 3 
Microscopic examination of feces, 149 

pathologic findings in, 150 

preparation of slides for, 149 
Milk curdling ferment, 136 

estimation of, 136 

method of Leo, 136 
Minims to cubic centimeters (table), 300 
Motor functions of stomach, 137 
iodoform test for, 138 
tests for, 137 

of intestines, 146 
Mucus in the feces, 157 
Myelocytes, 49 

N 
Nematodes, 113 
Nephelometer, the, 73 
Newmann's orcin test, 212 
Newton's rings, 37 
Nitrogenous equilibrium, 183 
Nucleo-albumin, 200 
Nylander's bismuth test, 203 

O 

Objectives, 3 
apochromatic, 5 
plane, 3 

oil-immersion, 5 
Oblique illumination, 6 
Ocular or eye-piece, 2 
compensating, 5 
finder in, 3 
Occult bleeding, 131, 153, 216 

acetic acid-ether-guaiac test, 132 
causes of, in stomach, 131 
in the urine, 216 
in the feces, 153 
-blood tests, 131, 154 
in feces, 153 

preparation of patient for, 153 
technic in feces, 154 
significance of, in feces, 154 
Weber's test, 131 
Office laboratory cabinet, 274 
Oliguria, 167 
Opsonic methods. 64-73 
bacterial emulsions, 66 
diagnosis of infections by, 71 
general considerations. 65 
immunization of mixed infections, 72 
method of obtaining index, 67 ' 
"pool"-serum, 67 
rules for treatment by, 68 
standardization of vaccines, 69 
the nephelometer, 73 
the serum, 67 
the technic, 66 
the theory, 64 
therapeutic application, 69 
washed white corpuscles, 67 
Organic acids in stomach, 129 

constituents of urine, 181 
Oxalic acid in the urine, 192 
Oxybutyric acid, 213 
Oxyuris vermicularis, 117 



Pappenheim's solution, 299 
method, 156 



INDEX. 



.",07 



Paramecium eoli. 107 
Pentoses in urine. 212 
Newmann'B test for, 212 

Penzoldts method. 137 
Peptie activity, 132 

Mitt's method of estimating, 133 

Sailer and Farr on, 135 
Peptone in stomach digestion, 131 
Percentage of red cells, 40 

method of estimating, 40 
Pericardial fluid. 238 
Peritoneal fluid. 236 

normal character of. 2: 1 >t> 

pathologic characteristics of, 236 
Peritonitis, blood-pressure in, 88 
Phenylhydrazin test. 204 
Phosphates in the urine, 176 

estimation of, 177 
Plasmodium malaria?, 96 

clinical relation of, 98 

detection of. 97 

varieties of, 96 
Platvhelminthes. 107 
Pleural fluid, 237 

microscopic examination of, 237, 238 

stained specimen of, 237 

the exudate, 237 

the transudate. 237 
Pneumococcus, 248 
Polymorphonuclear leukocytes, 49 
Polyuria, 167 

Portable urinalysis set, 275 
Postural variations in blood-pressure, 83 

summary of, 84 
Pressure of cerebrospinal fluid, 231 

pathologic variations in, 232 

physiologic variations in, 232 
Progressive pernicious anemia, 53 
Proteose, 200 
Protozoa, 103 

Pulmonary actinomycosis, 30 
Pulse pressure, 82 

factors influencing, 83 

method of determining, 83 
Purdy's test for albumin, 198 

quantitative method, 208 
Purin bases, 189 
Pus cells, 15 

in the urine, 217 
Pyuria, 217 

microscopic appearance of, 217 

significance of, 217 

tests for, 217 

Q 
Quantitative estimation of hydrochloric 
acid, 129 

R 
Reaction of the urine, 172 
Red cells, estimation of, 37 

centrifuge method, 39 

per cent , 40 

Thoma-Zeiss method, 37 

volume-index, 41 
Relapsing fever, spirochsete of, 99 
Renal disease, blood-pressure in, 86 
Rhizopodia, 103 

Robert's differential density method. 206 
Rontgen examination of stomach, 138 

Pfahler on, 139 

technic of, 141 
Russell and Brodie's coagulometer, 93 



Salicyluric acid te=t in urine, 146 
Schmidt method, 146 



Schmidt method, diet for, 146 

testing motor functions by, 148 
Serous stools, 143 
Serum-albumin test. 200 
Serum-globulin test, 200 
Shiga's bacillus, 156 
Specific gravity of the blood, ::i 

of the urine, 168 
Spectroscope, 57 

application of, 58 

description of, 57 

in blood examination, 57 
Spermaturia, 222 
Sphygmomanometer, the, 79, 88 

arm-band for, 80 

clinical value of, 87 

Dr. Faught's, 82 . 

hand-bellows for, 80 

in angina pectoris, 87 

in cardiac disease, 85 

in diabetes, 87 

in diphtheria, 88 

in exophthalmic goiter, 87 

in heart-muscle degeneration, 86 

in lead colic, 88 

in peritonitis, 88 

in renal disease, 86 

in typhoid fever, 88 

method of using, 79 
Spirochsete of relapsing fever, 99 

pallida, 59 
description of, 59 
obtaining specimen of, 60 
staining methods for, 60 
Sporozoa, 106 
Sputum, 19-30, 278 

absence of, 19 

accidental discoloration of, 23 

air-content of, 23 

amount of. 19 

black, 21, 22 

blood streaked, 22 

casts in, 27 

composition of, 20 

concretions in, 25 

consistency of, 20 

crystals in, 28 

currant-jelly, 21 

Curschmann's spirals in, 27 

Dietrich's plugs in, 25 

elastic fibers in, 28 

eosinophiles in, 27 

epithelia in, 26 

examination blank forms for, 278 

foreign bodies in, 25 

frothy, 21 

gross examination of, 25 

"heart-failure" cells in, 27 

hemorrhagic, 22 

microscopic examination of, 26 

mucopurulent, 20 

mucus in, 20 

numular, 21 

origin of, 19 

odor of, 24 

purulent, 21 

pus cells in, 26 

reaction of, 23 

red cells in, 27 

rusty, 21 

scanty, 19 

serous, 20 

shreds of tissue in, 21 

staining specimen of, 29 

tumor cells in, 29 

viscid, 20 



308 



INDEX. 



Sputum, yellow, 21, 22 
Stage, the warm, 9 
Staining methods, 45, 60, 297 
Ehrlich's triacid, 45 
eosin and methylene-blue, 45 
Giemsa, 60 

Oppenheim and Sachs', 60 
Romanowski, 46 
Wood's, 60 
Stains, 290-299 
carbol fuchsin, 296 
carbol gentian-violet, 290 
Chezinsky's, 290 
Ehrlich's, 290 
Gabbott's, 297 
Giemsa, 291 
Gram's, 297 
Hasting's, 295 
Hematoxylin-eosin, 290 
Loeffler's, 296 
Pappenheim's, 299 
Romanowski's, 294 
"Soloid," 292 
"Universal," 291 
Unna's orcin, 297 
Starch in test-meal, 131 

and sugar digestion, 137 
Sterilization, 255-262 
autoclave method of, 259 
chemical, 262 
dry heat, 256 

intermittent method of, 259 
steam, 257 
Sterilizer of Arnold, 261 
Stools, 143 

color of, 144 
Stomach, 118 
acids of digestion in, 126 
contents, 118 
alcohol in, 130 
blood in, 131 

detection of fatty acids in, 129 
detection of starch in, 137 
fatty acids in, 130 
hydrochloric acid in, 126 
method of obtaining specimen of, 118 
microscopic examination of, 131 
milk curdling ferment in, 136 
Penzoldt's method for, 137 
peptone in, 130 
starch in, 131 

test for hydrochloric acid in, 127 
test for lactic acid in, 130 
test for total acidity in, 128 
Daland-Faught gastric apparatus, 118 
determining capacity of, 125 
Ewald test-breakfast, 121 
inflation of, 123 
lavage of, 126 

modified Ewald breakfast, 121 
motor functions of, 137 
peptic activity of, 132 
Rontgen examination of, 138 
test-meal removal, 120 
vomitus, 118 

x-ray examination of, 138 
Streptococcus aerogenes, 155 
Strongyloides intestinalis, 113 
Substage condenser, 4 
Synovial fluid, 238 
Systolic blood-pressure, 80 
definition of, 82 
method of obtaining, 80 
recording, 83 

T 
Tables, 300, 301 
cubic centimeters to minims, 301 



Tables, grains to grams, 300 

grams to grains, 301 

minims to cubic centimeters, 300 
Tahret's test, 198 
Tape worms, 107 
Technic of gastric lavage, 126 
Tenia echinococcus, 112 

lanceolatus, 110 

lata, 109 

mediocanellata, 111 

nana, 111 

sagniata, 111 

solium, 110 

vulgaris, 110 
Teniidse, 110 

Terms used in hematology, 47 
Test-diet, 146 

period of passage of, 147 
Test-meal, 121 

technic of removal of, 122 
Tests for occult bleeding, 131 
Thoma-Zeiss hemocytometer, 36 
Thread worms, 113 
Tone of blood-vessels, 78 
Topfer's reagent, 127 
Total acidity of gastric contents, 123 

quantitative estimation of, 128 
Trapp's formula, 172 
Trematodes, 107 
Trichomonas intestinalis, 105 

pulmonale, 105 

vaginalis, 105 
Trichina spiralis, 115 

development of, 115 
Trichocephalus dispar, 114 
Trypanosomiasis, 100 
Trypanosomum gambiense, 100, 106 

staining of, 106 
Trommer's test, 204 
Tube casts in urine. 218 
Tubercle bacillus, 156, 244 

Czaplewsky's stain for, 247 

differentiation of, 246 

Gabbott's stain for, 247 

microscopic appearance of, 245 

Pappenheim's stain for, 247 

presence in feces, 156 

Rosenberger's technic for, 156 

staining methods, 246 
peculiarities of, 245 

Ziehl-Nielsen stain, 246 
Typhoid fever, 83, 94 

blood-pressure in, 88 

coagulation time in, 94 

Widal reaction in, 61 
Tyrosin, 193 

U 

Uffleman's test, 130 
Uncinaria Americana, 116 

duodenale, 115 
Urates, 189 

microscopic appearance of, 191 

qualitative tests for, 191 
Urea, 182 

detection of, 186 

estimation of, 186 

in disease, 185 

properties of, 185 
Uric acid', 187 

isolation of, 188 

microscopic appearance of, 188 

properties of, 188 

quantitative determination of, 189 

significance of, in urine, 188 

tests for, 188 
Urine, 16, 161-227 



INDEX. 



309 



Urine, abnormal constituents of, 194 
acetone in, 212 
albumin in. 194 
alkaline, 173 
alkapton in, 165 
amount of. 166 
anuria, 167 

average daily excretion, 166 
bacteria in, 16 
bile acids in, 215 
bile pigments in, 214 
Bird's formula, 172 
blood-pressure effecting, 88 
blood in, 215 

Cammidge reaction in, 210 
carbonates in, 224 
casts in, 218 

catheterized specimen of, 162 
chemical composition, 174 
chlorids in, 180-181 
cholesterin, 225 
chyle in, 222 
color of, 164 
concretions in, 225 
creatinin in, 191 
crystals in, 224 
cylindroids in, 221 
cystin in, 193, 225 
decomposition changes in, 162 
detection of phosphates in, 176, 177 
determination of acidity of, 174 

of alkalinity of, 174 
diacetic acid in, 213 
Diazo reaction in, 226 
directions for collecting, 161 
drugs and poisons in, 227 
earthy phosphates in, 178 
epithelia in, 218 

estimation of phosphates in, 177 
general characteristics of, 163 
glucose in, 201 
hemoglobin in, 217 
hippuric acid in, 191 
hydruria, 167 
increased acidity of, 173 
indican in, 179, 180 
inorganic constituents of, 175 

sediment of, 223 
lactose in, 212 
leucin in, 193 
maltose in, 212 
melanuria, 165 
method of collection of, 161 
Metz's formula, 172 
microscopic examination of, 193 
nitrogenous equilibrium of, 183 
odor of, 165 
oliguria, 167 
organic constituents of, 181 



Urine, oxalates in. 192 

oxybutyric acid in, 213 

passage of turbid, L63 

pentoses in, 212 

phosphates in, 176 

polariscopic examination of, 211 

polyuria, 167 

preparation of slide of, 224 

preservation of sample of, 163 

purin bases in, 199 

pyuria, 217 

reaction of, 172 

specific gravity of, 168 

spermaturia, 222 

sulphates in, 224 

sulphates, mineral in, 179 
preformed in, 179 

tests for melanin in, 165 

tests for phosphates in, 177 

test for reaction of, 172 

the sample of, 161 

Trapp's formula for, 172 

twenty-four hours' sample of, 162 

tyrosin in, 193 

urates in, 189 

urea in, 182 

uric acid in, 187 

urinometer for, 168 

urobilin in, 215 

Westphal balance for, 170 
Urinary sediments, 11 

microscopic search for, 12 

preparation of slide of, 11 
Urinalysis report blank, 277 
Urinometer, 168 
Urobilin, 215 

significance of, 215 

tests for, 215 

V 
Vernier scale, 9 
Volume-index of Russell, 41 

W 

Warm stage, 9 

Weber's test for occult bleeding, 121 

Westphal balance, 170 

White cells, estimation of, 42 

Widal reaction, 61 

cultures for, 62 

request for, 284 

serum dilutions for, 63 

serum for, 62 
Wright's coagulometer, 90 



X-ray examination or stomach, 138 
Pfahler on, 139 
technic of, 141 



■b n i%y 



^r 



